No. 1. Proteins: peptide bond, their detection.
Proteins are macromolecules of linear polyamides formed by a-amino acids as a result of a polycondensation reaction in biological objects.
Squirrels are macromolecular compounds built from amino acids. 20 amino acids are involved in making proteins. They link together into long chains that form the backbone of a large molecular weight protein molecule.
Functions of proteins in the body
The combination of peculiar chemical and physical properties of proteins provides this particular class of organic compounds with a central role in the phenomena of life.
Proteins have the following biological properties, or perform the following main functions in living organisms:
1. Catalytic function of proteins. All biological catalysts - enzymes are proteins. To date, thousands of enzymes have been characterized, many of them isolated in crystalline form. Almost all enzymes are powerful catalysts, increasing the rates of reactions by at least a million times. This function of proteins is unique, not characteristic of other polymeric molecules.
2. Nutritional (reserve function of proteins). These are, first of all, proteins intended for nutrition of the developing embryo: milk casein, egg ovalbumin, storage proteins of plant seeds. A number of other proteins are undoubtedly used in the body as a source of amino acids, which, in turn, are precursors of biologically active substances that regulate the metabolic process.
3. Transport function of proteins. Many small molecules and ions are transported by specific proteins. For example, the respiratory function of blood, namely the transport of oxygen, is performed by hemoglobin molecules, a protein in red blood cells. Serum albumins are involved in lipid transport. A number of other whey proteins form complexes with fats, copper, iron, thyroxine, vitamin A and other compounds, ensuring their delivery to the appropriate organs.
4. Protective function of proteins. The main function of protection is performed by the immunological system, which provides the synthesis of specific protective proteins - antibodies - in response to the entry of bacteria, toxins or viruses (antigens) into the body. Antibodies bind antigens, interacting with them, and thereby neutralize their biological effect and maintain the normal state of the body. The coagulation of a blood plasma protein - fibrinogen - and the formation of a blood clot that protects against blood loss during injuries is another example of the protective function of proteins.
5. Contractile function of proteins. Many proteins are involved in the act of muscle contraction and relaxation. The main role in these processes is played by actin and myosin - specific proteins of muscle tissue. The contractile function is also inherent in the proteins of subcellular structures, which provides the finest processes of cell vital activity,
6. Structural function of proteins. Proteins with this function rank first among other proteins in the human body. Structural proteins such as collagen are widely distributed in connective tissue; keratin in hair, nails, skin; elastin - in the vascular walls, etc.
7. Hormonal (regulatory) function of proteins. Metabolism in the body is regulated by various mechanisms. In this regulation, an important place is occupied by hormones produced by endocrine glands. A number of hormones are represented by proteins or polypeptides, for example, hormones of the pituitary gland, pancreas, etc.
Peptide bond
Formally, the formation of a protein macromolecule can be represented as a polycondensation reaction of α-amino acids.
From a chemical point of view, proteins are high-molecular nitrogen-containing organic compounds (polyamides), whose molecules are built from amino acid residues. Protein monomers are α-amino acids, common feature which is the presence of a carboxyl group -COOH and an amino group -NH 2 at the second carbon atom (α-carbon atom):
Based on the results of studying the products of protein hydrolysis and put forward by A.Ya. Danilevsky's ideas about the role of peptide bonds -CO-NH- in the construction of a protein molecule, the German scientist E. Fischer proposed at the beginning of the 20th century the peptide theory of the structure of proteins. According to this theory, proteins are linear polymers of α-amino acids linked by a peptide bond - polypeptides:
In each peptide, one terminal amino acid residue has a free α-amino group (N-terminus) and the other has a free α-carboxyl group (C-terminus). The structure of peptides is usually depicted starting from the N-terminal amino acid. In this case, amino acid residues are indicated by symbols. For example: Ala-Tyr-Leu-Ser-Tyr- - Cys. This entry denotes a peptide in which the N-terminal α-amino acid is lyatsya alanine, and the C-terminal - cysteine. When reading such a record, the endings of the names of all acids, except for the last ones, change to - "yl": alanyl-tyrosyl-leucyl-seryl-tyrosyl--cysteine. The length of the peptide chain in peptides and proteins found in the body ranges from two to hundreds and thousands of amino acid residues.
No. 2. Classification of simple proteins.
To simple (proteins) include proteins that, when hydrolyzed, give only amino acids.
Proteinoids ____simple proteins of animal origin, insoluble in water, salt solutions, dilute acids and alkalis. They perform mainly supporting functions (for example, collagen, keratin
protamines - positively charged nuclear proteins, with a molecular weight of 10-12 kDa. Approximately 80% are composed of alkaline amino acids, which makes it possible for them to interact with nucleic acids through ionic bonds. They take part in the regulation of gene activity. Well soluble in water;
histones - nuclear proteins that play an important role in the regulation of gene activity. They are found in all eukaryotic cells, and are divided into 5 classes, differing in molecular weight and amino acid. The molecular weight of histones is in the range from 11 to 22 kDa, and the differences in the amino acid composition relate to lysine and arginine, the content of which varies from 11 to 29% and from 2 to 14%, respectively;
prolamins - insoluble in water, but soluble in 70% alcohol, chemical structure features - a lot of proline, glutamic acid, no lysine ,
glutelins - soluble in alkaline solutions ,
globulins - proteins that are insoluble in water and in a semi-saturated solution of ammonium sulphate, but soluble in aqueous solutions of salts, alkalis and acids. Molecular weight - 90-100 kDa;
albumins - proteins of animal and plant tissues, soluble in water and saline solutions. The molecular weight is 69 kDa;
scleroproteins - proteins of the supporting tissues of animals
Examples of simple proteins are silk fibroin, egg serum albumin, pepsin, etc.
Number 3. Methods for isolation and precipitation (purification) of proteins.
No. 4. Proteins as polyelectrolytes. Isoelectric point of a protein.
Proteins are amphoteric polyelectrolytes, i.e. exhibit both acidic and basic properties. This is due to the presence in protein molecules of amino acid radicals capable of ionization, as well as free α-amino and α-carboxyl groups at the ends of peptide chains. Acidic properties of the protein are given by acidic amino acids (aspartic, glutamic), and alkaline properties - by basic amino acids (lysine, arginine, histidine).
The charge of a protein molecule depends on the ionization of acidic and basic groups of amino acid radicals. Depending on the ratio of negative and positive groups, the protein molecule as a whole acquires a total positive or negative charge. When a protein solution is acidified, the degree of ionization of anionic groups decreases, while that of cationic groups increases; when alkalized - vice versa. At a certain pH value, the number of positively and negatively charged groups becomes the same, and the isoelectric state of the protein appears (the total charge is 0). The pH value at which the protein is in the isoelectric state is called the isoelectric point and is denoted pI, similar to amino acids. For most proteins, pI lies in the range of 5.5-7.0, which indicates a certain predominance of acidic amino acids in proteins. However, there are also alkaline proteins, for example, salmin - the main protein from salmon milt (pl=12). In addition, there are proteins that have a very low pI value, for example, pepsin, an enzyme of gastric juice (pl=l). At the isoelectric point, proteins are very unstable and precipitate easily, having the least solubility.
If the protein is not in an isoelectric state, then in an electric field its molecules will move towards the cathode or anode, depending on the sign of the total charge and at a speed proportional to its value; this is the essence of the electrophoresis method. This method can separate proteins with different pI values.
Although proteins have buffer properties, their capacity at physiological pH values is limited. The exception is proteins containing a lot of histidine, since only the histidine radical has buffer properties in the pH range of 6-8. There are very few of these proteins. For example, hemoglobin, containing almost 8% histidine, is a powerful intracellular buffer in red blood cells, maintaining the pH of the blood at a constant level.
No. 5. Physico-chemical properties of proteins.
Proteins have different chemical, physical and biological properties, which are determined by the amino acid composition and spatial organization of each protein. The chemical reactions of proteins are very diverse, they are due to the presence of NH 2 -, COOH groups and radicals of various nature. These are reactions of nitration, acylation, alkylation, esterification, redox and others. Proteins have acid-base, buffer, colloidal and osmotic properties.
Acid-base properties of proteins
Chemical properties. With weak heating of aqueous solutions of proteins, denaturation occurs. This creates a precipitate.
When proteins are heated with acids, hydrolysis occurs, and a mixture of amino acids is formed.
Physico-chemical properties of proteins
Proteins have a high molecular weight.
The charge of a protein molecule. All proteins have at least one free -NH and -COOH group.
Protein solutions- colloidal solutions with different properties. Proteins are acidic and basic. Acidic proteins contain a lot of glu and asp, which have additional carboxyl and fewer amino groups. There are many lys and args in alkaline proteins. Each protein molecule in an aqueous solution is surrounded by a hydration shell, since proteins have many hydrophilic groups (-COOH, -OH, -NH 2, -SH) due to amino acids. In aqueous solutions, the protein molecule has a charge. The charge of protein in water can change depending on the pH.
Protein precipitation. Proteins have a hydration shell, a charge that prevents sticking. For deposition, it is necessary to remove the hydrate shell and charge.
1. Hydration. The process of hydration means the binding of water by proteins, while they exhibit hydrophilic properties: they swell, their mass and volume increase. Swelling of the protein is accompanied by its partial dissolution. The hydrophilicity of individual proteins depends on their structure. The hydrophilic amide (–CO–NH–, peptide bond), amine (NH2) and carboxyl (COOH) groups present in the composition and located on the surface of the protein macromolecule attract water molecules, strictly orienting them to the surface of the molecule. Surrounding the protein globules, the hydrate (water) shell prevents the stability of protein solutions. At the isoelectric point, proteins have the least ability to bind water, the hydration shell around the protein molecules is destroyed, so they combine to form large aggregates. Aggregation of protein molecules also occurs when they are dehydrated with some organic solvents, such as ethyl alcohol. This leads to the precipitation of proteins. When the pH of the medium changes, the protein macromolecule becomes charged, and its hydration capacity changes.
Precipitation reactions are divided into two types.
Salting out of proteins: (NH 4)SO 4 - only the hydration shell is removed, the protein retains all types of its structure, all bonds, retains its native properties. Such proteins can then be re-dissolved and used.
Precipitation with loss of native protein properties is an irreversible process. The hydration shell and charge are removed from the protein, various properties in the protein are violated. For example, salts of copper, mercury, arsenic, iron, concentrated inorganic acids - HNO 3 , H 2 SO 4 , HCl, organic acids, alkaloids - tannins, mercury iodide. The addition of organic solvents lowers the degree of hydration and leads to precipitation of the protein. Acetone is used as such solvent. Proteins are also precipitated with the help of salts, for example, ammonium sulfate. The principle of this method is based on the fact that with an increase in the salt concentration in the solution, the ionic atmospheres formed by the protein counterions are compressed, which contributes to their convergence to a critical distance, at which the intermolecular forces of van der Waals attraction outweigh the Coulomb forces of repulsion of the counterions. This leads to the adhesion of protein particles and their precipitation.
When boiling, protein molecules begin to move randomly, collide, the charge is removed, and the hydration shell decreases.
To detect proteins in solution, the following are used:
color reactions;
precipitation reactions.
Methods for isolation and purification of proteins.
salting out;
electrophoresis;
chromatography: adsorption, splitting;
ultracentrifugation.
homogenization- the cells are ground to a homogeneous mass;
extraction of proteins with water or water-salt solutions;
Structural organization of proteins.
Primary Structure- determined by the sequence of amino acids in the peptide chain, stabilized by covalent peptide bonds (insulin, pepsin, chymotrypsin).
secondary structure- spatial structure of the protein. This is either a spiral or a folding. Hydrogen bonds are created.
Tertiary structure globular and fibrillar proteins. They stabilize hydrogen bonds, electrostatic forces (COO-, NH3+), hydrophobic forces, sulfide bridges, are determined by the primary structure. Globular proteins - all enzymes, hemoglobin, myoglobin. Fibrillar proteins - collagen, myosin, actin.
Quaternary structure- found only in some proteins. Such proteins are built from several peptides. Each peptide has its own primary, secondary, tertiary structure, called protomers. Several protomers join together to form one molecule. One protomer does not function as a protein, but only in conjunction with other protomers.
Example: hemoglobin \u003d -globule + -globule - carries O 2 in the aggregate, and not separately.
Protein can renature. This requires a very short exposure to agents.
6) Methods for detecting proteins.
Proteins are high-molecular biological polymers, the structural (monomeric) units of which are -amino acids. Amino acids in proteins are linked to each other by peptide bonds. the formation of which occurs due to the carboxyl group standing at -carbon atom of one amino acid and -amine group of another amino acid with the release of a water molecule. The monomeric units of proteins are called amino acid residues.
Peptides, polypeptides and proteins differ not only in quantity, composition, but also in the sequence of amino acid residues, physicochemical properties and functions performed in the body. The molecular weight of proteins varies from 6 thousand to 1 million or more. The chemical and physical properties of proteins are due to the chemical nature and physico-chemical properties of the radicals that make up their amino acid residues. Methods for the detection and quantification of proteins in biological objects and food products, as well as their isolation from tissues and biological fluids, are based on physical and chemical properties these compounds.
Proteins when interacting with certain chemicals give colored compounds. The formation of these compounds occurs with the participation of amino acid radicals, their specific groups or peptide bonds. Color reactions allow you to set the presence of a protein in a biological object or solution and prove the presence certain amino acids in a protein molecule. On the basis of color reactions, some methods for the quantitative determination of proteins and amino acids have been developed.
Consider universal biuret and ninhydrin reactions, since all proteins give them. Xantoprotein reaction, Fohl reaction and others are specific, since they are due to the radical groups of certain amino acids in the protein molecule.
Color reactions allow you to establish the presence of a protein in the material under study and the presence of certain amino acids in its molecules.
Biuret reaction. The reaction is due to the presence in proteins, peptides, polypeptides peptide bonds, which in an alkaline medium form with copper(II) ions complex compounds colored in purple (with a red or blue tinge) color. The color is due to the presence of at least two groups in the molecule -CO-NH- connected directly to each other or with the participation of a carbon or nitrogen atom.
Copper (II) ions are connected by two ionic bonds with =C─O ˉ groups and four coordination bonds with nitrogen atoms (=N−).
The color intensity depends on the amount of protein in the solution. This makes it possible to use this reaction for the quantitative determination of protein. The color of the colored solutions depends on the length of the polypeptide chain. Proteins give a blue-violet color; the products of their hydrolysis (poly- and oligopeptides) are red or pink in color. The biuret reaction is given not only by proteins, peptides and polypeptides, but also by biuret (NH 2 -CO-NH-CO-NH 2), oxamide (NH 2 -CO-CO-NH 2), histidine.
The complex compound of copper (II) with peptide groups formed in an alkaline medium has the following structure:
Ninhydrin reaction. In this reaction, solutions of protein, polypeptides, peptides and free α-amino acids, when heated with ninhydrin, give a blue, blue-violet or pink-violet color. The color in this reaction develops due to the α-amino group.
-amino acids react very easily with ninhydrin. Along with them, Rueman's blue-violet is also formed by proteins, peptides, primary amines, ammonia, and some other compounds. Secondary amines such as proline and hydroxyproline give a yellow color.
The ninhydrin reaction is widely used to detect and quantify amino acids.
xantoprotein reaction. This reaction indicates the presence of aromatic amino acid residues in proteins - tyrosine, phenylalanine, tryptophan. It is based on the nitration of the benzene ring of the radicals of these amino acids with the formation of yellow-colored nitro compounds (Greek "Xanthos" - yellow). Using tyrosine as an example, this reaction can be described in the form of the following equations.
In an alkaline environment, nitro derivatives of amino acids form salts of the quinoid structure, colored orange. The xantoprotein reaction is produced by benzene and its homologues, phenol and other aromatic compounds.
Reactions to amino acids containing a thiol group in a reduced or oxidized state (cysteine, cystine).
Fol's reaction. When boiling with alkali, sulfur is easily cleaved off from cysteine in the form of hydrogen sulfide, which in an alkaline medium forms sodium sulfide:
In this regard, the reactions for determining thiol-containing amino acids in solution are divided into two stages:
The transition of sulfur from organic to inorganic state
Detection of sulfur in solution
To detect sodium sulfide, lead acetate is used, which, when interacting with sodium hydroxide, turns into its plumbite:
Pb(CH 3 COO) 2 + 2NaOH Pb(ONa) 2 +2CH 3 COOH
As a result of the interaction of sulfur ions and lead, black or brown lead sulfide is formed:
Na 2 S + Pb(ONa) 2 + 2 H 2 O PbS (black sediment) + 4NaOH
To determine sulfur-containing amino acids, an equal volume of sodium hydroxide and a few drops of lead acetate solution are added to the test solution. With intensive boiling for 3-5 minutes, the liquid turns black.
The presence of cystine can be determined using this reaction, since cystine is easily reduced to cysteine.
Millon reaction:
This is a reaction to the amino acid tyrosine.
Free phenolic hydroxyls of tyrosine molecules, when interacting with salts, give compounds of the mercury salt of the nitro derivative of tyrosine, colored pinkish red:
Pauli reaction for histidine and tyrosine . The Pauli reaction makes it possible to detect the amino acids histidine and tyrosine in the protein, which form cherry-red complex compounds with diazobenzenesulfonic acid. Diazobenzenesulfonic acid is formed in the reaction of diazotization when sulfanilic acid reacts with sodium nitrite in an acidic medium:
An equal volume of an acidic solution of sulfanilic acid (prepared using hydrochloric acid) and a double volume of sodium nitrite solution are added to the test solution, mixed thoroughly and soda (sodium carbonate) is immediately added. After stirring, the mixture turns cherry red, provided that histidine or tyrosine is present in the test solution.
Adamkevich-Hopkins-Kohl (Schulz-Raspail) reaction to tryptophan (reaction to the indole group). Tryptophan reacts in an acidic environment with aldehydes, forming colored condensation products. The reaction proceeds due to the interaction of the indole ring of tryptophan with aldehyde. It is known that formaldehyde is formed from glyoxylic acid in the presence of sulfuric acid:
R 
Solutions containing tryptophan in the presence of glyoxylic and sulfuric acids give a red-violet color.
Glyoxylic acid is always present in small amounts in glacial acetic acid. Therefore, the reaction can be carried out using acetic acid. At the same time, an equal volume of glacial (concentrated) acetic acid is added to the test solution and gently heated until the precipitate dissolves. After cooling, a volume of concentrated sulfuric acid equal to the added volume of glyoxylic acid is added to the mixture carefully along the wall (to avoid mixing liquids). After 5-10 minutes, the formation of a red-violet ring is observed at the interface between the two layers. If you mix the layers, the contents of the dish will evenly turn purple.
To 
condensation of tryptophan with formaldehyde:
The condensation product is oxidized to bis-2-tryptophanylcarbinol, which in the presence of mineral acids forms blue-violet salts:
7) Classification of proteins. Methods for studying the amino acid composition.
Strict nomenclature and classification of proteins still does not exist. The names of proteins are given randomly, most often taking into account the source of protein isolation or taking into account its solubility in certain solvents, the shape of the molecule, etc.
Proteins are classified according to composition, particle shape, solubility, amino acid composition, origin, etc.
1. Composition Proteins are divided into two large groups: simple and complex proteins.
Simple (proteins) include proteins that give only amino acids upon hydrolysis (proteinoids, protamines, histones, prolamins, glutelins, globulins, albumins). Examples of simple proteins are silk fibroin, egg serum albumin, pepsin, etc.
Complex (proteids) include proteins composed of a simple protein and an additional (prosthetic) group of non-protein nature. The group of complex proteins is divided into several subgroups depending on the nature of the non-protein component:
Metalloproteins containing in their composition metals (Fe, Cu, Mg, etc.) associated directly with the polypeptide chain;
Phosphoproteins - contain residues of phosphoric acid, which are attached to the protein molecule by ester bonds at the site of the hydroxyl groups of serine, threonine;
Glycoproteins - their prosthetic groups are carbohydrates;
Chromoproteins - consist of a simple protein and a colored non-protein compound associated with it, all chromoproteins are biologically very active; as prosthetic groups, they may contain derivatives of porphyrin, isoalloxazine, and carotene;
Lipoproteins - prosthetic group lipids - triglycerides (fats) and phosphatides;
Nucleoproteins are proteins that consist of a single protein and a nucleic acid linked to it. These proteins play a colossal role in the life of the body and will be discussed below. They are part of any cell, some nucleoproteins exist in nature in the form of special particles with pathogenic activity (viruses).
2. Particle shape- proteins are divided into fibrillar (thread-like) and globular (spherical) (see page 30).
3. By solubility and characteristics of the amino acid composition the following groups of simple proteins are distinguished:
Proteinoids - proteins of supporting tissues (bones, cartilage, ligaments, tendons, hair, nails, skin, etc.). These are mainly fibrillar proteins with a large molecular weight (> 150,000 Da), insoluble in common solvents: water, salt and water-alcohol mixtures. They dissolve only in specific solvents;
Protamines (the simplest proteins) - proteins that are soluble in water and contain 80-90% arginine and a limited set (6-8) of other amino acids, are present in the milk of various fish. Due to the high content of arginine, they have basic properties, their molecular weight is relatively small and is approximately equal to 4000-12000 Da. They are a protein component in the composition of nucleoproteins;
Histones are highly soluble in water and dilute solutions of acids (0.1 N), have a high content of amino acids: arginine, lysine and histidine (at least 30%) and therefore have basic properties. These proteins are found in significant amounts in the nuclei of cells as part of nucleoproteins and play an important role in the regulation of nucleic acid metabolism. The molecular weight of histones is small and equal to 11000-24000 Da;
Globulins are proteins that are insoluble in water and saline solutions with a salt concentration of more than 7%. Globulins are completely precipitated at 50% saturation of the solution with ammonium sulfate. These proteins are characterized by a high content of glycine (3.5%), their molecular weight > 100,000 Da. Globulins are weakly acidic or neutral proteins (p1=6-7.3);
Albumins are proteins that are highly soluble in water and strong saline solutions, and the salt concentration (NH 4) 2 S0 4 should not exceed 50% of saturation. At higher concentrations, albumins are salted out. Compared to globulins, these proteins contain three times less glycine and have a molecular weight of 40,000-70,000 Da. Albumins have an excess negative charge and acidic properties (pl=4.7) due to the high content of glutamic acid;
Prolamins are a group of plant proteins found in the gluten of cereals. They are soluble only in 60-80% aqueous solution of ethyl alcohol. Prolamins have a characteristic amino acid composition: they contain a lot (20-50%) of glutamic acid and proline (10-15%), which is why they got their name. Their molecular weight is over 100,000 Da;
Glutelins - vegetable proteins are insoluble in water, salt solutions and ethanol, but soluble in dilute (0.1 N) solutions of alkalis and acids. In terms of amino acid composition and molecular weight, they are similar to prolamins, but contain more arginine and less proline.
Methods for studying the amino acid composition
Proteins are broken down into amino acids by enzymes in the digestive juices. Two important conclusions were made: 1) proteins contain amino acids; 2) methods of hydrolysis can be used to study the chemical, in particular amino acid, composition of proteins.
To study the amino acid composition of proteins, a combination of acidic (HCl), alkaline [Ba (OH) 2] and, less often, enzymatic hydrolysis or one of them. It has been established that during the hydrolysis of a pure protein that does not contain impurities, 20 different α-amino acids are released. All other amino acids discovered in the tissues of animals, plants and microorganisms (more than 300) exist in nature in a free state or in the form of short peptides or complexes with other organic substances.
The first step in determining the primary structure of proteins is the qualitative and quantitative assessment of the amino acid composition of a given individual protein. It must be remembered that for the study you need to have a certain amount of pure protein, without impurities of other proteins or peptides.
Acid hydrolysis of protein
To determine the amino acid composition, it is necessary to destroy all peptide bonds in the protein. The analyzed protein is hydrolyzed in 6 mol/l HC1 at a temperature of about 110 °C for 24 hours. As a result of this treatment, peptide bonds in the protein are destroyed, and only free amino acids are present in the hydrolyzate. In addition, glutamine and asparagine are hydrolyzed to glutamic and aspartic acids (i.e., the amide bond in the radical is broken and the amino group is cleaved off from them).
Separation of amino acids using ion exchange chromatography
The mixture of amino acids obtained by acid hydrolysis of proteins is separated in a column with a cation exchange resin. Such a synthetic resin contains negatively charged groups (for example, sulfonic acid residues -SO 3 -) strongly associated with it, to which Na + ions are attached (Fig. 1-4).
A mixture of amino acids is introduced into the cation exchanger in an acidic environment (pH 3.0), where the amino acids are mainly cations, i. carry a positive charge. Positively charged amino acids attach to negatively charged resin particles. The greater the total charge of the amino acid, the stronger its bond with the resin. Thus, the amino acids lysine, arginine, and histidine bind most strongly to the cation exchanger, while aspartic and glutamic acids bind the most weakly.
The release of amino acids from the column is carried out by eluting (eluting) them with a buffer solution with increasing ionic strength (ie, with increasing NaCl concentration) and pH. With an increase in pH, amino acids lose a proton, as a result, their positive charge decreases, and hence the bond strength with negatively charged resin particles.
Each amino acid exits the column at a specific pH and ionic strength. By collecting the solution (eluate) from the lower end of the column in the form of small portions, fractions containing individual amino acids can be obtained.
(for more details on "hydrolysis" see question #10)
8) Chemical bonds in the protein structure.
9) The concept of the hierarchy and structural organization of proteins. (see question #12)
10) Protein hydrolysis. Reaction chemistry (stepping, catalysts, reagents, reaction conditions) - a complete description of hydrolysis.
11) Chemical transformations of proteins.
Denaturation and renaturation
When protein solutions are heated to 60-80% or under the action of reagents that destroy non-covalent bonds in proteins, the tertiary (quaternary) and secondary structure of the protein molecule is destroyed, it takes the form of a random random coil to a greater or lesser extent. This process is called denaturation. Acids, alkalis, alcohols, phenols, urea, guanidine chloride, etc. can be used as denaturing reagents. The essence of their action is that they form hydrogen bonds with = NH and = CO - groups of the peptide backbone and with acid groups of amino acid radicals, replacing their own intramolecular hydrogen bonds in the protein, as a result of which the secondary and tertiary structures change. During denaturation, the solubility of the protein decreases, it “coagulates” (for example, when boiling a chicken egg), and the biological activity of the protein is lost. Based on this, for example, the use of an aqueous solution of carbolic acid (phenol) as an antiseptic. Under certain conditions, with slow cooling of a solution of a denatured protein, renaturation occurs - the restoration of the original (native) conformation. This confirms the fact that the nature of the folding of the peptide chain is determined by the primary structure.
The process of denaturation of an individual protein molecule, leading to the disintegration of its "rigid" three-dimensional structure, is sometimes called the melting of the molecule. Almost any noticeable change in external conditions, such as heating or a significant change in pH, leads to a consistent violation of the quaternary, tertiary and secondary structures of the protein. Usually, denaturation is caused by an increase in temperature, the action of strong acids and alkalis, salts of heavy metals, certain solvents (alcohol), radiation, etc.
Denaturation often leads to the process of aggregation of protein particles into larger ones in a colloidal solution of protein molecules. Visually, this looks, for example, as the formation of a "protein" when frying eggs.
Renaturation is the reverse process of denaturation, in which proteins return to their natural structure. It should be noted that not all proteins are able to renature; in most proteins, denaturation is irreversible. If, during protein denaturation, physicochemical changes are associated with the transition of the polypeptide chain from a densely packed (ordered) state to a disordered state, then during renaturation, the ability of proteins to self-organize is manifested, the path of which is predetermined by the sequence of amino acids in the polypeptide chain, that is, its primary structure determined by hereditary information . In living cells, this information is probably decisive for the transformation of a disordered polypeptide chain during or after its biosynthesis on the ribosome into the structure of a native protein molecule. When double-stranded DNA molecules are heated to a temperature of about 100 ° C, the hydrogen bonds between the bases are broken, and the complementary strands diverge - the DNA denatures. However, upon slow cooling, the complementary strands can reconnect into a regular double helix. This ability of DNA to renature is used to produce artificial DNA hybrid molecules.
Natural protein bodies are endowed with a certain, strictly defined spatial configuration and have a number of characteristic physicochemical and biological properties at physiological temperatures and pH values. Under the influence of various physical and chemical factors, proteins undergo coagulation and precipitate, losing their native properties. Thus, denaturation should be understood as a violation of the general plan of the unique structure of the native protein molecule, mainly its tertiary structure, leading to the loss of its characteristic properties (solubility, electrophoretic mobility, biological activity, etc.). Most proteins denature when their solutions are heated above 50–60°C.
External manifestations of denaturation are reduced to a loss of solubility, especially at the isoelectric point, an increase in the viscosity of protein solutions, an increase in the number of free functional SH-groups, and a change in the nature of X-ray scattering. The most characteristic sign of denaturation is a sharp decrease or complete loss by the protein of its biological activity (catalytic, antigenic or hormonal). During protein denaturation caused by 8M urea or another agent, mostly non-covalent bonds (in particular, hydrophobic interactions and hydrogen bonds) are destroyed. Disulfide bonds are broken in the presence of the reducing agent mercaptoethanol, while the peptide bonds of the backbone of the polypeptide chain itself are not affected. Under these conditions, globules of native protein molecules unfold and random and disordered structures are formed (Fig.)
Denaturation of a protein molecule (scheme).
a - initial state; b - beginning reversible violation of the molecular structure; c - irreversible deployment of the polypeptide chain.
Denaturation and renaturation of ribonuclease (according to Anfinsen).
a - deployment (urea + mercaptoethanol); b - refolding.
1. Protein hydrolysis: H+
[− NH2─CH─ CO─NH─CH─CO − ]n +2nH2O → n NH2 − CH − COOH + n NH2 ─ CH ─ COOH
│ │ │ │
Amino acid 1 amino acid 2
2. Precipitation of proteins:
a) reversible
Protein in solution ↔ protein precipitate. Occurs under the action of solutions of salts Na+, K+
b) irreversible (denaturation)
During denaturation under the influence of external factors (temperature; mechanical action - pressure, rubbing, shaking, ultrasound; the action of chemical agents - acids, alkalis, etc.), a change occurs in the secondary, tertiary and quaternary structures of the protein macromolecule, i.e. its native spatial structure. The primary structure, and, consequently, the chemical composition of the protein does not change.
During denaturation, the physical properties of proteins change: solubility decreases, biological activity is lost. At the same time, the activity of some chemical groups increases, the effect of proteolytic enzymes on proteins is facilitated, and, consequently, it is more easily hydrolyzed.
For example, albumin - egg white - at a temperature of 60-70 ° is precipitated from a solution (coagulates), losing the ability to dissolve in water.
Scheme of the process of protein denaturation (destruction of the tertiary and secondary structures of protein molecules)
3. Burning proteins
Proteins burn with the formation of nitrogen, carbon dioxide, water, and some other substances. Burning is accompanied by the characteristic smell of burnt feathers.
4. Color (qualitative) reactions to proteins:
a) xantoprotein reaction (for amino acid residues containing benzene rings):
Protein + HNO3 (conc.) → yellow color
b) biuret reaction (for peptide bonds):
Protein + CuSO4 (sat) + NaOH (conc) → bright purple color
c) cysteine reaction (for amino acid residues containing sulfur):
Protein + NaOH + Pb(CH3COO)2 → Black staining
Proteins are the basis of all life on Earth and perform various functions in organisms.
Salting out proteins
Salting out is the process of isolating proteins from aqueous solutions with neutral solutions of concentrated salts of alkali and alkaline earth metals. When high concentrations of salts are added to the protein solution, the dehydration of the protein particles and the removal of the charge occur, while the proteins precipitate. The degree of protein precipitation depends on the ionic strength of the precipitant solution, the size of the particles of the protein molecule, the magnitude of its charge, and hydrophilicity. Different proteins precipitate at different salt concentrations. Therefore, in sediments obtained by gradually increasing the concentration of salts, individual proteins are in different fractions. Salting out of proteins is a reversible process, and after the salt is removed, the protein regains its natural properties. Therefore, salting out is used in clinical practice in the separation of blood serum proteins, as well as in the isolation and purification of various proteins.
Added anions and cations destroy the hydrated protein shell of proteins, which is one of the stability factors of protein solutions. Most often, solutions of Na and ammonium sulfates are used. Many proteins differ in the size of the hydration shell and the magnitude of the charge. Each protein has its own salting out zone. After removal of the salting out agent, the protein retains its biological activity and physiochemical properties. In clinical practice, the salting out method is used to separate globulins (with the addition of 50% ammonium sulfate (NH4)2SO4 a precipitate precipitates) and albumins (with the addition of 100% ammonium sulfate (NH4)2SO4 a precipitate precipitates).
Salting out is influenced by:
1) nature and concentration of salt;
2) pH environments;
3) temperature.
The main role is played by the valencies of the ions.
12) Features of the organization of the primary, secondary, tertiary structure of the protein.
At present, the existence of four levels has been experimentally proven structural organization protein molecule: primary, secondary, tertiary and quaternary structure.
Squirrels- high-molecular organic compounds, consisting of residues of α-amino acids.
AT protein composition includes carbon, hydrogen, nitrogen, oxygen, sulfur. Some proteins form complexes with other molecules containing phosphorus, iron, zinc and copper.
Proteins have a large molecular weight: egg albumin - 36,000, hemoglobin - 152,000, myosin - 500,000. For comparison: the molecular weight of alcohol is 46, acetic acid - 60, benzene - 78.
Amino acid composition of proteins
Squirrels- non-periodic polymers, the monomers of which are α-amino acids. Usually, 20 types of α-amino acids are called protein monomers, although more than 170 of them have been found in cells and tissues.
Depending on whether amino acids can be synthesized in the body of humans and other animals, there are: nonessential amino acids- can be synthesized; essential amino acids- cannot be synthesized. Essential amino acids must be ingested with food. Plants synthesize all kinds of amino acids.
Depending on the amino acid composition, proteins are: complete- contain the entire set of amino acids; defective- some amino acids are absent in their composition. If proteins are made up of only amino acids, they are called simple. If proteins contain, in addition to amino acids, also a non-amino acid component (a prosthetic group), they are called complex. The prosthetic group can be represented by metals (metalloproteins), carbohydrates (glycoproteins), lipids (lipoproteins), nucleic acids (nucleoproteins).
All amino acids contain: 1) a carboxyl group (-COOH), 2) an amino group (-NH 2), 3) a radical or R-group (the rest of the molecule). The structure of the radical different types amino acids are different. Depending on the number of amino groups and carboxyl groups that make up amino acids, there are: neutral amino acids having one carboxyl group and one amino group; basic amino acids having more than one amino group; acidic amino acids having more than one carboxyl group.
Amino acids are amphoteric compounds, since in solution they can act as both acids and bases. In aqueous solutions, amino acids exist in different ionic forms.
Peptide bond
Peptides — organic matter, consisting of amino acid residues connected by a peptide bond.
The formation of peptides occurs as a result of the condensation reaction of amino acids. When the amino group of one amino acid interacts with the carboxyl group of another, a covalent nitrogen-carbon bond arises between them, which is called peptide. Depending on the number of amino acid residues that make up the peptide, there are dipeptides, tripeptides, tetrapeptides etc. The formation of a peptide bond can be repeated many times. This leads to the formation polypeptides. At one end of the peptide there is a free amino group (it is called the N-terminus), and at the other end there is a free carboxyl group (it is called the C-terminus).
Spatial organization of protein molecules
The performance of certain specific functions by proteins depends on the spatial configuration of their molecules, in addition, it is energetically unfavorable for the cell to keep proteins in an expanded form, in the form of a chain, therefore, polypeptide chains undergo folding, acquiring a certain three-dimensional structure, or conformation. Allocate 4 levels spatial organization of proteins.
Primary structure of a protein- the sequence of amino acid residues in the polypeptide chain that makes up the protein molecule. The bond between amino acids is peptide.
If a protein molecule consists of only 10 amino acid residues, then the number theoretically options protein molecules that differ in the order of alternation of amino acids - 10 20. With 20 amino acids, you can make even more diverse combinations of them. About ten thousand different proteins have been found in the human body, which differ both from each other and from the proteins of other organisms.
It is the primary structure of the protein molecule that determines the properties of the protein molecules and its spatial configuration. The replacement of just one amino acid for another in the polypeptide chain leads to a change in the properties and functions of the protein. For example, the replacement of the sixth glutamine amino acid in the β-subunit of hemoglobin with valine leads to the fact that the hemoglobin molecule as a whole cannot perform its main function - oxygen transport; in such cases, a person develops a disease - sickle cell anemia.
secondary structure- ordered folding of the polypeptide chain into a spiral (looks like a stretched spring). The coils of the helix are strengthened by hydrogen bonds between carboxyl groups and amino groups. Almost all CO and NH groups take part in the formation of hydrogen bonds. They are weaker than peptide ones, but, repeating many times, they impart stability and rigidity to this configuration. At the level of the secondary structure, there are proteins: fibroin (silk, web), keratin (hair, nails), collagen (tendons).
Tertiary structure- packing of polypeptide chains into globules, resulting from the occurrence of chemical bonds (hydrogen, ionic, disulfide) and the establishment of hydrophobic interactions between radicals of amino acid residues. The main role in the formation of the tertiary structure is played by hydrophilic-hydrophobic interactions. In aqueous solutions, hydrophobic radicals tend to hide from water, grouping inside the globule, while hydrophilic radicals, as a result of hydration (interaction with water dipoles), tend to appear on the surface of the molecule. In some proteins, the tertiary structure is stabilized by disulfide covalent bonds that form between the sulfur atoms of the two cysteine residues. At the level of the tertiary structure, there are enzymes, antibodies, some hormones.
Quaternary structure characteristic of complex proteins, the molecules of which are formed by two or more globules. Subunits are held in the molecule by ionic, hydrophobic, and electrostatic interactions. Sometimes, during the formation of a quaternary structure, disulfide bonds occur between subunits. The most studied protein with a quaternary structure is hemoglobin. It is formed by two α-subunits (141 amino acid residues) and two β-subunits (146 amino acid residues). Each subunit is associated with a heme molecule containing iron.
If for some reason the spatial conformation of proteins deviates from normal, the protein cannot perform its functions. For example, the cause of "mad cow disease" (spongiform encephalopathy) is an abnormal conformation of prions, the surface proteins of nerve cells.
Protein properties
The amino acid composition, the structure of the protein molecule determine its properties. Proteins combine essential and acid properties determined by amino acid radicals: the more acidic amino acids in a protein, the more pronounced its acidic properties. The ability to give and attach H + determine buffer properties of proteins; one of the most powerful buffers is hemoglobin in erythrocytes, which maintains the pH of the blood at a constant level. There are soluble proteins (fibrinogen), there are insoluble proteins that perform mechanical functions (fibroin, keratin, collagen). There are chemically active proteins (enzymes), there are chemically inactive, resistant to various environmental conditions and extremely unstable.
External factors (heat, ultraviolet radiation, heavy metals and their salts, pH changes, radiation, dehydration)
can cause a violation of the structural organization of the protein molecule. The process of losing the three-dimensional conformation inherent in a given protein molecule is called denaturation. The cause of denaturation is the breaking of bonds that stabilize a particular protein structure. Initially, the weakest ties are torn, and when conditions become tougher, even stronger ones. Therefore, first the quaternary, then the tertiary and secondary structures are lost. A change in the spatial configuration leads to a change in the properties of the protein and, as a result, makes it impossible for the protein to perform its inherent biological functions. If denaturation is not accompanied by the destruction of the primary structure, then it can be reversible, in this case, self-healing of the conformation characteristic of the protein occurs. Such denaturation is subjected, for example, to membrane receptor proteins. The process of restoring the structure of a protein after denaturation is called renaturation. If the restoration of the spatial configuration of the protein is impossible, then denaturation is called irreversible.
Functions of proteins
| Function | Examples and explanations |
|---|---|
| Construction | Proteins are involved in the formation of cellular and extracellular structures: they are part of cell membranes (lipoproteins, glycoproteins), hair (keratin), tendons (collagen), etc. |
| Transport | The blood protein hemoglobin attaches oxygen and transports it from the lungs to all tissues and organs, and from them carbon dioxide transfers to the lungs; The composition of cell membranes includes special proteins that provide an active and strictly selective transfer of certain substances and ions from the cell to the external environment and vice versa. |
| Regulatory | Protein hormones are involved in the regulation of metabolic processes. For example, the hormone insulin regulates blood glucose levels, promotes glycogen synthesis, and increases the formation of fats from carbohydrates. |
| Protective | In response to the penetration of foreign proteins or microorganisms (antigens) into the body, special proteins are formed - antibodies that can bind and neutralize them. Fibrin, formed from fibrinogen, helps to stop bleeding. |
| Motor | The contractile proteins actin and myosin provide muscle contraction in multicellular animals. |
| Signal | Molecules of proteins are embedded in the surface membrane of the cell, capable of changing their tertiary structure in response to the action of environmental factors, thus receiving signals from the external environment and transmitting commands to the cell. |
| Reserve | In the body of animals, proteins, as a rule, are not stored, with the exception of egg albumin, milk casein. But thanks to proteins in the body, some substances can be stored in reserve, for example, during the breakdown of hemoglobin, iron is not excreted from the body, but is stored, forming a complex with the ferritin protein. |
| Energy | With the breakdown of 1 g of protein to the final products, 17.6 kJ is released. First, proteins break down into amino acids, and then to the end products - water, carbon dioxide and ammonia. However, proteins are used as an energy source only when other sources (carbohydrates and fats) are used up. |
| catalytic | One of the most important functions of proteins. Provided with proteins - enzymes that accelerate bio chemical reactions occurring in cells. For example, ribulose biphosphate carboxylase catalyzes CO2 fixation during photosynthesis. |
Enzymes
Enzymes, or enzymes, is a special class of proteins that are biological catalysts. Thanks to enzymes, biochemical reactions proceed at a tremendous speed. The rate of enzymatic reactions is tens of thousands of times (and sometimes millions) higher than the rate of reactions involving inorganic catalysts. The substance on which an enzyme acts is called substrate.
Enzymes are globular proteins structural features Enzymes can be divided into two groups: simple and complex. simple enzymes are simple proteins, i.e. consist only of amino acids. Complex enzymes are complex proteins, i.e. in addition to the protein part, they include a group of non-protein nature - cofactor. For some enzymes, vitamins act as cofactors. In the enzyme molecule, a special part is isolated, called the active center. active center- a small region of the enzyme (from three to twelve amino acid residues), where the binding of the substrate or substrates occurs with the formation of an enzyme-substrate complex. Upon completion of the reaction, the enzyme-substrate complex decomposes into an enzyme and a reaction product(s). Some enzymes have (other than active) allosteric centers- sites to which regulators of the rate of enzyme work are attached ( allosteric enzymes).
Enzymatic catalysis reactions are characterized by: 1) high efficiency, 2) strict selectivity and direction of action, 3) substrate specificity, 4) fine and precise regulation. The substrate and reaction specificity of enzymatic catalysis reactions is explained by the hypotheses of E. Fischer (1890) and D. Koshland (1959).
E. Fisher (key-lock hypothesis) suggested that the spatial configurations of the active site of the enzyme and the substrate should correspond exactly to each other. The substrate is compared to the "key", the enzyme - to the "lock".
D. Koshland (hypothesis "hand-glove") suggested that the spatial correspondence between the structure of the substrate and the active center of the enzyme is created only at the moment of their interaction with each other. This hypothesis is also called induced fit hypothesis.
The rate of enzymatic reactions depends on: 1) temperature, 2) enzyme concentration, 3) substrate concentration, 4) pH. It should be emphasized that since enzymes are proteins, their activity is highest under physiologically normal conditions.
Most enzymes can only work at temperatures between 0 and 40°C. Within these limits, the reaction rate increases by about 2 times for every 10 °C rise in temperature. At temperatures above 40 °C, the protein undergoes denaturation and the activity of the enzyme decreases. At temperatures close to freezing, the enzymes are inactivated.
With an increase in the amount of substrate, the rate of the enzymatic reaction increases until the number of substrate molecules becomes equal to the number of enzyme molecules. With a further increase in the amount of substrate, the rate will not increase, since the active sites of the enzyme are saturated. An increase in the enzyme concentration leads to an increase in catalytic activity, since a larger number of substrate molecules undergo transformations per unit time.
For each enzyme, there is an optimal pH value at which it exhibits maximum activity (pepsin - 2.0, salivary amylase - 6.8, pancreatic lipase - 9.0). At higher or lower pH values, the activity of the enzyme decreases. With sharp shifts in pH, the enzyme denatures.
The speed of allosteric enzymes is regulated by substances that attach to allosteric centers. If these substances speed up the reaction, they are called activators if they slow down - inhibitors.
Enzyme classification
According to the type of catalyzed chemical transformations, enzymes are divided into 6 classes:
- oxidoreductase(transfer of hydrogen, oxygen or electron atoms from one substance to another - dehydrogenase),
- transferase(transfer of a methyl, acyl, phosphate or amino group from one substance to another - transaminase),
- hydrolases(hydrolysis reactions in which two products are formed from the substrate - amylase, lipase),
- lyases(non-hydrolytic addition to the substrate or cleavage of a group of atoms from it, while C-C, C-N, C-O, C-S bonds can be broken - decarboxylase),
- isomerase(intramolecular rearrangement - isomerase),
- ligases(connection of two molecules as a result of the formation C-C connections, C-N, C-O, C-S - synthetase).
Classes are in turn subdivided into subclasses and subsubclasses. In the current international classification, each enzyme has a specific code, consisting of four numbers separated by dots. The first number is the class, the second is the subclass, the third is the subclass, the fourth is the serial number of the enzyme in this subclass, for example, the arginase code is 3.5.3.1.
Go to lectures number 2"The structure and functions of carbohydrates and lipids"
Go to lectures №4"The structure and functions of ATP nucleic acids"
Before talking about the properties of proteins, it is worth giving a brief definition of this concept. These are high-molecular organic substances that consist of alpha-amino acids connected by a peptide bond. Proteins are an important part of human and animal nutrition, since not all amino acids are produced by the body - some come from food. What are their properties and functions?
Amphoteric
This is the first feature of proteins. Amphoteric refers to their ability to exhibit both acidic and basic properties.
Proteins in their structure have several types of chemical groups that are able to ionize in a solution of H 2 O. These include:
- carboxyl residues. Glutamic and aspartic acids, to be precise.
- nitrogen containing groups.ε-amino group of lysine, arginine residue CNH(NH 2) and imidazole residue of a heterocyclic alpha-amino acid called histidine.
Each protein has such a feature as an isoelectric point. This concept is understood as the acidity of the medium at which the surface or molecule does not have an electric charge. Under such conditions, protein hydration and solubility are minimized.
The indicator is determined by the ratio of basic and acidic amino acid residues. In the first case, the point falls on the alkaline region. In the second - sour.
Solubility
According to this property, proteins are divided into a small classification. Here's what they are:
- Soluble. They are called albumins. They are sparingly soluble in concentrated saline solutions and coagulate when heated. This reaction is called denaturation. The molecular weight of albumins is about 65,000. They do not contain carbohydrates. And substances that consist of albumin are called albuminoids. These include egg white, plant seeds and blood serum.
- insoluble. They are called scleroproteins. A striking example is keratin, a fibrillar protein with mechanical strength second only to chitin. It is from this substance that nails, hair, the ramphotheque of bird beaks and feathers, as well as rhinoceros horns, are composed. This group of proteins also includes cytokeratins. This is the structural material of intracellular filaments of the cytoskeleton of epithelial cells. Another insoluble protein is a fibrillar protein called fibroin.
- hydrophilic. They actively interact with water and absorb it. These include proteins of the intercellular substance, nucleus and cytoplasm. Including the notorious fibroin and keratin.
- hydrophobic. They repel water. These include proteins that are components of biological membranes.
Denaturation
This is the name of the process of modification of a protein molecule under the influence of certain destabilizing factors. The amino acid sequence remains the same. But proteins lose their natural properties (hydrophilicity, solubility, and others).
It should be noted that any significant change in external conditions can lead to violations of protein structures. Most often, denaturation is provoked by an increase in temperature, as well as the effect of alkali, strong acid, radiation, heavy metal salts, and even certain solvents on the protein.
Interestingly, often denaturation leads to the fact that protein particles are aggregated into larger ones. A prime example is scrambled eggs. After all, everyone is familiar with how, in the process of frying, the protein is formed from a transparent liquid.
You should also talk about such a phenomenon as renaturation. This process is the reverse of denaturation. During it, proteins return to their natural structure. And it's really possible. A group of chemists from the US and Australia have found a way to renature a hard-boiled egg. It will only take a few minutes. And this will require urea (diamide of carbonic acid) and centrifugation.
Structure
It must be said separately, since we are talking about the importance of proteins. In total, there are four levels of structural organization:
- Primary. The sequence of amino acid residues in a polypeptide chain is meant. The main feature is conservative motives. These are stable combinations of amino acid residues. They are found in many complex and simple proteins.
- Secondary. This refers to the ordering of some local fragment of the polypeptide chain, which is stabilized by hydrogen bonds.
- Tertiary. This is the spatial structure of the polypeptide chain. This level consists of some secondary elements (they are stabilized different types interactions, where hydrophobic are the most important). Here, ionic, hydrogen, covalent bonds are involved in stabilization.
- Quaternary. It is also called domain or subunit. This level comprises relative position chains of polypeptides as part of a whole protein complex. It is interesting that proteins with a quaternary structure include not only identical, but also different chains of polypeptides.
This division was proposed by a Danish biochemist named K. Lindstrom-Lang. And even if it is considered obsolete, they still continue to use it.
Building types
Speaking about the properties of proteins, it should also be noted that these substances are divided into three groups in accordance with the type of structure. Namely:
- fibrillar proteins. They have a filamentous elongated structure and a large molecular weight. Most of them are insoluble in water. The structure of these proteins is stabilized by interactions between polypeptide chains (they consist of at least two amino acid residues). It is the fibrillar substances that form the polymer, fibrils, microtubules and microfilaments.
- globular proteins. The type of structure determines their solubility in water. And the general shape of the molecule is spherical.
- membrane proteins. The structure of these substances is interesting feature. They have domains that cross the cell membrane, but parts of them protrude into the cytoplasm and extracellular environment. These proteins play the role of receptors - they transmit signals and are responsible for the transmembrane transport of nutrients. It is important to note that they are very specific. Each protein passes only a certain molecule or signal.
Simple
You can also tell a little more about them. Simple proteins consist only of chains of polypeptides. These include:
- Protamine. Nuclear low molecular weight protein. Its presence is the protection of DNA from the action of nucleases - enzymes that attack nucleic acids.
- Histones. Strongly basic simple proteins. They are concentrated in the nuclei of plant and animal cells. They take part in the "packaging" of DNA strands in the nucleus, and also in processes such as repair, replication and transcription.
- Albumins. They have already been mentioned above. The most famous albumins are serum and egg.
- Globulin. Participates in blood clotting, as well as in other immune reactions.
- Prolamins. These are storage proteins of cereals. Their names are always different. In wheat, they are called ptyalins. Barley has hordeins. Oats have avsnins. Interestingly, prolamins are divided into their own classes of proteins. There are only two of them: S-rich (with sulfur content) and S-poor (without it).
Complex
What about complex proteins? They contain prosthetic groups or those without amino acids. These include:
- Glycoproteins. They contain carbohydrate residues with a covalent bond. These complex proteins are the most important structural component of cell membranes. They also include many hormones. And the glycoproteins of erythrocyte membranes determine the blood type.
- Lipoproteins. They consist of lipids (fat-like substances) and play the role of "transport" of these substances in the blood.
- Metalloproteins. These proteins in the body are of great importance, since without them the exchange of iron does not proceed. Their molecules contain metal ions. And typical representatives of this class are transferrin, hemosiderin and ferritin.
- Nucleoproteins. They consist of RKN and DNA that do not have a covalent bond. A prominent representative is chromatin. It is in its composition that genetic information is realized, DNA is repaired and replicated.
- Phosphoproteins. They are covalently bonded phosphoric acid residues. An example is casein, which is originally found in milk as a calcium salt (in bound form).
- Chromoproteins. They have a simple structure: a protein and a colored component belonging to the prosthetic group. They take part in cellular respiration, photosynthesis, redox reactions, etc. Also, without chromoproteins, energy accumulation does not occur.
Metabolism
Much has already been said above about the physicochemical properties of proteins. Their role in metabolism should also be mentioned.
There are amino acids that are indispensable because they are not synthesized by living organisms. Mammals themselves get them from food. In the process of digestion, protein is destroyed. This process begins with denaturation when it is placed in an acidic environment. Then - hydrolysis, in which enzymes participate.
Certain amino acids that the body eventually receives are involved in the process of protein synthesis, the properties of which are necessary for its full existence. And the rest is processed into glucose - a monosaccharide, which is one of the main sources of energy. Protein is very important in terms of diets or starvation. If it does not come with food, the body will begin to "eat itself" - process its own proteins, especially muscle proteins.
Biosynthesis
Considering the physicochemical properties of proteins, it is necessary to focus on such a topic as biosynthesis. These substances are formed on the basis of the information that is encoded in the genes. Any protein is a unique sequence of amino acid residues determined by the gene encoding it.
How does this happen? A gene encoding a protein transfers information from DNA to RNA. This is called transcription. In most cases, synthesis then occurs on ribosomes - this is the most important organelle of a living cell. This process is called translation.
There is also the so-called non-ribosomal synthesis. It is also worth mentioning, since we are talking about the importance of proteins. This type of synthesis is observed in some bacteria and lower fungi. The process is carried out through a high molecular weight protein complex (known as NRS synthase), and ribosomes do not take part in this.
And, of course, there is also chemical synthesis. It can be used to synthesize short proteins. For this, methods such as chemical ligation are used. This is the opposite of the notorious biosynthesis on ribosomes. The same method can be used to obtain inhibitors of certain enzymes.
In addition, thanks to chemical synthesis, it is possible to introduce into the composition of proteins those amino acid residues that are not found in ordinary substances. Let's say those whose side chains have fluorescent labels.
It is worth mentioning that the methods of chemical synthesis are not perfect. There are certain restrictions. If the protein contains more than 300 residues, then the artificially synthesized substance is likely to receive an incorrect structure. And this will affect the properties.
Substances of animal origin
Their consideration should be given special attention. Animal protein is a substance found in eggs, meat, dairy products, poultry, seafood, and fish. They contain all the amino acids needed by the body, including 9 essential ones. Here are a number of the most important functions that animal protein performs:
- Catalysis of many chemical reactions. This substance launches them and accelerates them. Enzymatic proteins are “responsible” for this. If the body does not receive enough of them, then oxidation and reduction, connection and rupture molecular bonds, as well as the transportation of substances will not proceed fully. Interestingly, only a small part of the amino acids enter into various kinds of interactions. And an even smaller amount (3-4 residues) is directly involved in catalysis. All enzymes are divided into six classes - oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases. Each of them is responsible for a particular reaction.
- Formation of the cytoskeleton that forms the structure of cells.
- Immune, chemical and physical protection.
- Transportation of important components necessary for the growth and development of cells.
- The transmission of electrical impulses that are important for the functioning of the whole organism, since without them the interaction of cells is impossible.
And this is not all possible functions. But even so, the significance of these substances is clear. Protein synthesis in cells and in the body is impossible if a person does not eat its sources. And they are turkey meat, beef, lamb, rabbit meat. A lot of protein is found in eggs, sour cream, yogurt, cottage cheese, milk. You can also activate protein synthesis in the cells of the body by adding ham, offal, sausage, stew and veal to your diet.
The purpose of the lesson: to form the concept of a protein, its structure, physical and chemical properties.
During the classes
I. Organizational moment
II. Knowledge update
(Students are invited to repeat the topic "Amino acids" in advance.)
Two students work at the blackboard.
Exercise 1. Write the formulas for 2-aminopropanoic acid (alanine) and 3-methyl-2-aminobutanoic acid (valine). What other names for these acids can you suggest?
Task 2. Write the formula of 2-aminoethanoic acid. What other names for this acid do you know? Make a dipeptide from two residues of this acid. Specify the location of the peptide bond.
Frontal conversation.
What are the two functional groups in amino acids?
– What are amino acids in terms of acid-base properties? Due to what functional groups these properties are realized?
Give the concept of a peptide bond.
Can amino acids form hydrogen bonds? Due to what groups of atoms?
What substances are called polymers? Give examples of polymers known to you.
III. Setting a cognitive task
Students who worked at the blackboard report on the completed task.
The board shows a dipeptide consisting of two glycine residues, and the formulas of two amino acids: alanine and valine.
Can a dipeptide be formed from amino acids of different composition? (Slide 1.) In order to answer this question, pay attention to the place of the peptide bond in the dipeptide.
Answer. The amino group of one amino acid and the carboxyl group of another amino acid take part in the formation of a peptide bond; side radicals of amino acids are not involved in the formation of the dipeptide.
Is it possible to further attach amino acids to this substance? Justify the answer.
Answer. Accession is possible, because the dipeptide molecule has a free carboxyl group (C-terminus) and an amino group (N-terminus). The chain can grow on both sides (slide 2).
How many connection options can you offer?
Answer. Two. When the amino acid glycine is in first place and when the amino acid glycine is in second place (slide 3).
Answer. Proteins are linear biological polymers composed of amino acids.
Record this definition on your worksheets.
Here are two polypeptide chains. Which of the peptides can be part of the protein and why? (Slide 4.)
Answer. The first is because it is formed by α-amino acids.
What bonds form the primary structure of a protein?
Answer. The primary structure is formed by peptide bonds.
Record this in the table on your worksheet.
But a protein is a much more complex macromolecule than a linear polypeptide chain. In addition to the primary structure of the protein, it is necessary to consider secondary, tertiary, and, in some cases, quaternary structures. Hydrogen bonds play an important role in the formation of the secondary structure of a protein. Hydrogen bonds are formed by electronegative atoms (oxygen, nitrogen, etc.), with one of which a hydrogen atom is bonded, and all three atoms are on the same straight line.
Some proteins form a quaternary structure, which is also carried out due to hydrogen bonds, hydrophilic-hydrophobic interactions, and electrostatic forces of attraction. Some proteins with a quaternary structure consist of a metal ion and a protein part formed by several protein chains (different or identical in primary structure) (slide 7). Write on worksheets.
Proteins perform their functions correctly only in the presence of the appropriate tertiary (and quaternary, if any) structures.
Physical properties of proteins
Proteins are macromolecular compounds, i.e. These are substances with a high molecular weight. The molecular weight of proteins ranges from 5 thousand to millions of amu. (insulin - 6500 Da; influenza virus protein - 32 million Da).
The solubility of proteins in water depends on their functions. Molecules of fibrillar proteins are elongated, filamentous and tend to group one next to the other with the formation of fibers. This is the main building material for tendon, muscle and integumentary tissues. These proteins are insoluble in water.
The strength of protein molecules is simply amazing! Human hair is stronger than copper and can compete with special steels. A bundle of hair with an area of 1 cm 2 can withstand a weight of 5 tons, and on a female braid of 200 thousand hairs, you can lift a loaded KamAZ weighing 20 tons.
Globular proteins are folded into balls. In the body, they perform a number of biological functions that require their mobility. Therefore, globular proteins are soluble in water or in solutions of salts, acids or bases. Due to the large size of the molecules, solutions are formed, called colloidal. ( Demonstration of the dissolution of albumin in water.)
Chemical properties of proteins
Proteins are involved in not quite ordinary chemical reactions, tk. they are polymer molecules. Look at your work cards and answer the following questions.
Which bond is stronger: peptide or hydrogen?
Answer. Peptide, because this bond refers to a covalent chemical bond.
Which protein structures will be destroyed faster and easier?
Answer. Quaternary (if any), tertiary and secondary. The primary structure will last longer than others, because. it is formed by stronger bonds.
Denaturation is the destruction of a protein to its primary structure, i.e. peptide bonds are preserved (slide 8).
Demonstration of experience. Pour 4 ml of albumin solution into 5 small test tubes. Heat the first tube for 6–10 s (until cloudy). Add 2 ml of 3M HCl to the second tube. In the third - 2 ml of 3M NaOH. In the fourth - 5 drops of 0.1 M AgNO 3. In the fifth - 5 drops of 0.1 M NaNO 3.
After conducting the experiment, students fill in the gaps in the definition of the concept of "denaturation" on the worksheets.
Will proteins show their specific properties after denaturation?
Answer. Most proteins lose their activity during denaturation, tk. proteins show their specific properties only in the presence of tertiary and quaternary structures.
Do you think it is possible to destroy the primary structure of a protein?
Answer. Can. It occurs in the body when protein is digested.
One of the most important properties of proteins is their ability to hydrolyze. During protein hydrolysis, the primary structure is destroyed.
What substances are formed during the complete hydrolysis of a protein?
Answer. -amino acids.
Demonstration of experience (laid before the lesson). 2 ml of chicken protein solution are poured into two test tubes, 1 ml of a saturated solution of festal is added to one of them (the tablet is previously freed from the smooth shell). Festal is an enzyme preparation that facilitates digestion, which includes lipase (breaks down fats), amylase (breaks down carbohydrates), protease (breaks down proteins). Both test tubes are placed in a water bath at a temperature of 37–40 °C. Within 30 minutes, the process of "digestion" of the protein continues. At the end of heating, 2 ml of a saturated solution of ammonium sulfate or any other reagent that causes protein denaturation is added to both test tubes. In the first test tube (control), an abundant white precipitate is formed - the protein denatures. In the second test tube (experiment) such phenomena are not observed - protein hydrolysis occurred, and amino acids and peptides with a small molecular weight do not coagulate.
Based on the results of the experiment, fill in the gaps in the definition of "hydrolysis" on the worksheets.
What is the importance of protein hydrolysis for our body and where does it occur?
Answer. Obtaining amino acids for the needs of the body as a result of digestion processes begins in the stomach, ends in the duodenum.
Color reactions - qualitative reactions to proteins:
a) biuret reaction ( demonstration of experience);
b) xantoprotein reaction ( demonstration of experience).
Fill in the worksheets (pay attention to the conditions for the occurrence of these reactions, this will be needed for experiments in the next lesson).
WorksheetTopic: “Squirrels. Structure and properties»Proteins ________________________________________________________________________ Types of protein structures
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In the first half of the 19th century many chemists, and among them primarily J. von Liebig, gradually came to the conclusion that proteins are a special class of nitrogenous compounds. The name "proteins" (from the Greek.
protos the first) was proposed in 1840 by the Dutch chemist G. Mulder. PHYSICAL PROPERTIES Proteins are white in the solid state, but colorless in solution, unless they carry some chromophore (colored) group, such as hemoglobin. The solubility in water of different proteins varies greatly. It also varies with pH and with the concentration of salts in the solution, so that one can choose the conditions under which one protein will selectively precipitate in the presence of other proteins. This "salting out" method is widely used to isolate and purify proteins. The purified protein often precipitates out of solution as crystals.In comparison with other compounds, the molecular weight of proteins is very large - from several thousand to many millions of daltons. Therefore, during ultracentrifugation, proteins are precipitated, and, moreover, at different rates. Due to the presence of positively and negatively charged groups in protein molecules, they move at different speeds in an electric field. This is the basis of electrophoresis, a method used to isolate individual proteins from complex mixtures. Purification of proteins is also carried out by chromatography.
CHEMICAL PROPERTIES Structure. Proteins are polymers, i.e. molecules built like chains from repeating monomeric units, or subunits, the role of which they play a -amino acids. General formula amino acids where R a hydrogen atom or some organic group.A protein molecule (polypeptide chain) may consist of only a relatively small number of amino acids or several thousand monomer units. The connection of amino acids in the chain is possible because each of them has two different chemical groups: an amino group with basic properties,
NH2 , and an acidic carboxyl group, COOH. Both of these groups are affiliated with a - a carbon atom. The carboxyl group of one amino acid can form an amide (peptide) bond with the amino group of another amino acid:
After two amino acids have been connected in this way, the chain can be extended by adding a third to the second amino acid, and so on. As can be seen from the above equation, when a peptide bond is formed, a water molecule is released. In the presence of acids, alkalis or proteolytic enzymes, the reaction proceeds in the opposite direction: the polypeptide chain is cleaved into amino acids with the addition of water. This reaction is called hydrolysis. Hydrolysis proceeds spontaneously, and energy is required to combine amino acids into a polypeptide chain. A carboxyl group and an amide group (or a similar imide group in the case of the amino acid proline) are present in all amino acids, but the differences between amino acids are determined by the nature of that group, or "side chain", which is indicated above by the letter
R . The role of the side chain can be played by one hydrogen atom, as in the amino acid glycine, or by some bulky group, as in histidine and tryptophan. Some side chains are chemically inert, while others are highly reactive.Many thousands of different amino acids can be synthesized, and many different amino acids occur in nature, but only 20 types of amino acids are used for protein synthesis: alanine, arginine, asparagine, aspartic acid, valine, histidine, glycine, glutamine, glutamic acid, isoleucine, leucine, lysine , methionine, proline, serine, tyrosine, threonine, tryptophan, phenylalanine, and cysteine (in proteins, cysteine may be present as a dimer
cystine). True, there are other amino acids in some proteins, in addition to the regularly occurring twenty, but they are formed as a result of modification of any of the twenty listed after it has been included in the protein.optical activity. All amino acids, with the exception of glycine, a The carbon atom has four different groups attached. In terms of geometry, four different groups can be attached in two ways, and accordingly there are two possible configurations, or two isomers, related to each other as an object to its mirror image, i.e. like left hand to right. One configuration is called left, or left-handed ( L ), and the other right, or dextrorotatory ( D ), since two such isomers differ in the direction of rotation of the plane of polarized light. Only found in proteins L -amino acids (the exception is glycine; it can be represented by only one form, since two of its four groups are the same), and they all have optical activity (since there is only one isomer). D -amino acids are rare in nature; they are found in some antibiotics and the cell wall of bacteria.The sequence of amino acids. Amino acids in the polypeptide chain are not randomly arranged, but in a certain fixed order, and it is this order that determines the functions and properties of the protein. By varying the order of the 20 types of amino acids, you can get a huge number of different proteins, just like you can make up many different texts from the letters of the alphabet.In the past, determining the amino acid sequence of a protein often took several years. Direct determination is still a rather laborious task, although devices have been created that allow it to be carried out automatically. It is usually easier to determine the nucleotide sequence of the corresponding gene and derive the amino acid sequence of the protein from it. To date, the amino acid sequences of many hundreds of proteins have already been determined. The functions of decoded proteins are usually known, and this helps to imagine the possible functions of similar proteins formed, for example, in malignant neoplasms.
Complex proteins. Proteins consisting of only amino acids are called simple. Often, however, a metal atom or some chemical compound that is not an amino acid is attached to the polypeptide chain. Such proteins are called complex. An example is hemoglobin: it contains iron porphyrin, which gives it its red color and allows it to act as an oxygen carrier.The names of most complex proteins contain an indication of the nature of the attached groups: sugars are present in glycoproteins, fats in lipoproteins. If the catalytic activity of the enzyme depends on the attached group, then it is called a prosthetic group. Often, some vitamin plays the role of a prosthetic group or is part of it. Vitamin A, for example, attached to one of the proteins of the retina, determines its sensitivity to light.
Tertiary structure. What is important is not so much the amino acid sequence of the protein (primary structure), but the way it is laid in space. Along the entire length of the polypeptide chain, hydrogen ions form regular hydrogen bonds, which give it the shape of a spiral or layer (secondary structure). From the combination of such helices and layers, a compact form of the next order arises - the tertiary structure of the protein. Around the bonds that hold the monomeric links of the chain, rotations through small angles are possible. Therefore, from a purely geometric point of view, the number of possible configurations for any polypeptide chain is infinitely large. In reality, each protein normally exists in only one configuration, determined by its amino acid sequence. This structure is not rigid, it is, as it were, « breathes” fluctuates around a certain average configuration. The chain is folded into a configuration in which the free energy (the ability to do work) is minimal, just as a released spring is compressed only to a state corresponding to a minimum of free energy. Often, one part of the chain is rigidly linked to another disulfide ( SS) bonds between two cysteine residues. This is partly why cysteine among amino acids plays a particularly important role.The complexity of the structure of proteins is so great that it is not yet possible to calculate the tertiary structure of a protein, even if its amino acid sequence is known. But if it is possible to obtain protein crystals, then its tertiary structure can be determined by X-ray diffraction.
In structural, contractile, and some other proteins, the chains are elongated and several slightly folded chains lying side by side form fibrils; fibrils, in turn, fold into larger formations fibers. However, most proteins in solution are globular: the chains are coiled in a globule, like yarn in a ball. Free energy in this configuration is minimal, since the hydrophobic (“water-repellent”) amino acids are hidden inside the globule, and the hydrophilic (“water-attracting”) amino acids are on its surface.
Many proteins are complexes of several polypeptide chains. This structure is called the quaternary structure of the protein. The hemoglobin molecule, for example, is made up of four subunits, each of which is a globular protein.
Structural proteins, due to their linear configuration, form fibers in which the tensile strength is very high, while the globular configuration allows proteins to enter into specific interactions with other compounds. On the surface of the globule, with the correct laying of chains, cavities of a certain shape appear, in which reactive chemical groups are located. If a given protein is an enzyme, then another, usually smaller, molecule of some substance enters such a cavity, just as a key enters a lock; in this case, the configuration of the electron cloud of the molecule changes under the influence of chemical groups located in the cavity, and this forces it to react in a certain way. In this way, the enzyme catalyzes the reaction. Antibody molecules also have cavities in which various foreign substances bind and are thereby rendered harmless. The "key and lock" model, which explains the interaction of proteins with other compounds, makes it possible to understand the specificity of enzymes and antibodies, i.e. their ability to react only with certain compounds.
Proteins in different types of organisms. Proteins that perform the same function in different plant and animal species and therefore bear the same name also have a similar configuration. They, however, differ somewhat in their amino acid sequence. As species diverge from a common ancestor, some amino acids in certain positions are replaced by mutations with others. Harmful mutations that cause hereditary diseases are discarded by natural selection, but beneficial or at least neutral ones can be preserved. The closer two biological species are to each other, the less differences are found in their proteins.Some proteins change relatively quickly, others are quite conservative. The latter include, for example, cytochrome With a respiratory enzyme found in most living organisms. In humans and chimpanzees, its amino acid sequences are identical, and in cytochrome With wheat, only 38% of amino acids turned out to be different. Even comparing humans and bacteria, the similarity of cytochromes With(the differences affect 65% of the amino acids here) can still be seen, although the common ancestor of bacteria and humans lived on Earth about two billion years ago. Nowadays, comparison of amino acid sequences is often used to build a phylogenetic (genealogical) tree that reflects the evolutionary relationships between different organisms.
Denaturation. The synthesized protein molecule, folding, acquires its own configuration. This configuration, however, can be destroyed by heating, by changing the pH, by the action of organic solvents, and even by simply agitating the solution until bubbles appear on its surface. A protein altered in this way is called denatured; it loses its biological activity and usually becomes insoluble. Well-known examples of denatured protein are boiled eggs or whipped cream. Small proteins, containing only about a hundred amino acids, are able to renature, i.e. reacquire the original configuration. But most of the proteins are simply transformed into a mass of tangled polypeptide chains and do not restore their previous configuration.One of the main difficulties in isolating active proteins is their extreme sensitivity to denaturation. This property of proteins finds useful application in the preservation of food products: high temperature irreversibly denatures the enzymes of microorganisms, and the microorganisms die.
PROTEIN SYNTHESIS For protein synthesis, a living organism must have a system of enzymes capable of attaching one amino acid to another. A source of information is also needed that would determine which amino acids should be connected. Since there are thousands of types of proteins in the body, and each of them consists of an average of several hundred amino acids, the information required must be truly enormous. It is stored (similar to how a record is stored on a magnetic tape) in the nucleic acid molecules that make up genes. Cm . also HEREDITY; NUCLEIC ACIDS.Enzyme activation. A polypeptide chain synthesized from amino acids is not always a protein in its final form. Many enzymes are first synthesized as inactive precursors and become active only after another enzyme removes a few amino acids from one end of the chain. Some of the digestive enzymes, such as trypsin, are synthesized in this inactive form; these enzymes are activated in the digestive tract as a result of the removal of the terminal fragment of the chain. The hormone insulin, whose molecule in its active form consists of two short chains, is synthesized in the form of a single chain, the so-called. proinsulin. Then the middle part of this chain is removed, and the remaining fragments bind to each other, forming the active hormone molecule. Complex proteins are formed only after a certain chemical group is attached to the protein, and this attachment often also requires an enzyme.Metabolic circulation. After feeding an animal with amino acids labeled with radioactive isotopes of carbon, nitrogen or hydrogen, the label is quickly incorporated into its proteins. If labeled amino acids cease to enter the body, then the amount of label in proteins begins to decrease. These experiments show that the resulting proteins are not stored in the body until the end of life. All of them, with a few exceptions, are in a dynamic state, constantly decomposing to amino acids, and then re-synthesized.Some proteins break down when cells die and are destroyed. This happens all the time, for example, with red blood cells and epithelial cells lining the inner surface of the intestine. In addition, the breakdown and resynthesis of proteins also occur in living cells. Oddly enough, less is known about the breakdown of proteins than about their synthesis. What is clear, however, is that proteolytic enzymes are involved in the breakdown, similar to those that break down proteins into amino acids in the digestive tract.
The half-life of different proteins is different from several hours to many months. The only exception is the collagen molecule. Once formed, they remain stable and are not renewed or replaced. Over time, however, some of their properties, in particular elasticity, change, and since they are not renewed, certain age-related changes are the result of this, for example, the appearance of wrinkles on the skin.
synthetic proteins. Chemists have long since learned how to polymerize amino acids, but the amino acids are combined randomly, so that the products of such polymerization bear little resemblance to natural ones. True, it is possible to combine amino acids in a given order, which makes it possible to obtain some biologically active proteins, in particular insulin. The process is quite complicated, and in this way it is possible to obtain only those proteins whose molecules contain about a hundred amino acids. It is preferable instead to synthesize or isolate the nucleotide sequence of the gene corresponding to the desired amino acid sequence, and then introduce this gene into a bacterium, which will produce by replication a large amount of the desired product. This method, however, also has its drawbacks. Cm . See also GENETIC ENGINEERING. PROTEINS AND NUTRITION When proteins in the body are broken down into amino acids, these amino acids can be reused for protein synthesis. At the same time, the amino acids themselves are subject to decay, so that they are not fully utilized. It is also clear that during growth, pregnancy, and wound healing, protein synthesis must exceed degradation. The body continuously loses some proteins; these are the proteins of hair, nails and the surface layer of the skin. Therefore, for the synthesis of proteins, each organism must receive amino acids from food. Green plants are synthesized from CO 2 , water and ammonia or nitrates are all 20 amino acids found in proteins. Many bacteria are also able to synthesize amino acids in the presence of sugar (or some equivalent) and fixed nitrogen, but sugar is ultimately supplied by green plants. In animals, the ability to synthesize amino acids is limited; they obtain amino acids by eating green plants or other animals. In the digestive tract, the absorbed proteins are broken down into amino acids, the latter are absorbed, and the proteins characteristic of the given organism are built from them. None of the absorbed protein is incorporated into body structures as such. The only exception is that in many mammals, part of maternal antibodies can pass intact through the placenta into the fetal circulation, and through mother's milk (especially in ruminants) be transferred to the newborn immediately after birth.Need for proteins. It is clear that in order to maintain life, the body must receive a certain amount of protein from food. However, the size of this need depends on a number of factors. The body needs food both as a source of energy (calories) and as a material for building its structures. In the first place is the need for energy. This means that when there are few carbohydrates and fats in the diet, dietary proteins are used not for the synthesis of their own proteins, but as a source of calories. With prolonged fasting, even your own proteins are spent to meet energy needs. If there are enough carbohydrates in the diet, then protein intake can be reduced.nitrogen balance. On average approx. 16% of the total protein mass is nitrogen. When the amino acids that make up proteins are broken down, the nitrogen contained in them is excreted from the body in the urine and (to a lesser extent) in the feces in the form of various nitrogenous compounds. Therefore, it is convenient to use such an indicator as nitrogen balance to assess the quality of protein nutrition, i.e. the difference (in grams) between the amount of nitrogen taken into the body and the amount of nitrogen excreted per day. With normal nutrition in an adult, these amounts are equal. In a growing organism, the amount of excreted nitrogen is less than the amount of incoming, i.e. the balance is positive. With a lack of protein in the diet, the balance is negative. If there are enough calories in the diet, but the proteins are completely absent in it, the body saves proteins. At the same time, protein metabolism slows down, and the re-utilization of amino acids in protein synthesis proceeds as efficiently as possible. However, losses are inevitable, and nitrogenous compounds are still excreted in the urine and partly in the feces. The amount of nitrogen excreted from the body per day during protein starvation can serve as a measure of the daily lack of protein. It is natural to assume that by introducing into the diet an amount of protein equivalent to this deficiency, it is possible to restore the nitrogen balance. However, it is not. Having received this amount of protein, the body begins to use amino acids less efficiently, so some additional protein is required to restore the nitrogen balance.If the amount of protein in the diet exceeds what is necessary to maintain nitrogen balance, then there seems to be no harm from this. Excess amino acids are simply used as a source of energy. A particularly striking example is the Eskimo, who consume little carbohydrate and about ten times more protein than is required to maintain nitrogen balance. In most cases, however, using protein as an energy source is not beneficial, since you can get many more calories from a given amount of carbohydrates than from the same amount of protein. In poor countries, the population receives the necessary calories from carbohydrates and consumes a minimum amount of protein.
If the body receives the required number of calories in the form of non-protein foods, then the minimum amount of protein that maintains the nitrogen balance is approx. 30 g per day. Approximately as much protein is contained in four slices of bread or 0.5 liters of milk. A slightly larger amount is usually considered optimal; recommended from 50 to 70 g.
Essential amino acids. Until now, protein has been considered as a whole. Meanwhile, in order for protein synthesis to take place, all the necessary amino acids must be present in the body. Some of the amino acids the body of the animal itself is able to synthesize. They are called replaceable because they do not have to be present in the diet, it is only important that, in general, the intake of protein as a source of nitrogen is sufficient; then, with a shortage of non-essential amino acids, the body can synthesize them at the expense of those that are present in excess. The remaining "essential" amino acids cannot be synthesized and must be ingested with food. Essential for humans are valine, leucine, isoleucine, threonine, methionine, phenylalanine, tryptophan, histidine, lysine, and arginine. (Although arginine can be synthesized in the body, it is considered an essential amino acid because newborns and growing children produce insufficient amounts of it. On the other hand, for a person of mature age, the intake of some of these amino acids from food may become optional.)This list of essential amino acids is approximately the same in other vertebrates and even in insects. The nutritional value of proteins is usually determined by feeding them to growing rats and monitoring the weight gain of the animals.
The nutritional value of proteins. The nutritional value of a protein is determined by the essential amino acid that is most deficient. Let's illustrate this with an example. The proteins of our body contain an average of approx. 2% tryptophan (by weight). Let's say that the diet includes 10 g of protein containing 1% tryptophan, and that there are enough other essential amino acids in it. In our case, 10 g of this defective protein is essentially equivalent to 5 g of a complete one; the remaining 5 g can only serve as a source of energy. Note that, since amino acids are practically not stored in the body, and in order for protein synthesis to take place, all amino acids must be present simultaneously, the effect of the intake of essential amino acids can be detected only if they all enter the body at the same time.. The average composition of most animal proteins is close to the average composition of proteins in the human body, so we are unlikely to face amino acid deficiency if our diet is rich in foods such as meat, eggs, milk and cheese. However, there are proteins, such as gelatin (a product of collagen denaturation), which contain very few essential amino acids. Vegetable proteins, although they are better than gelatin in this sense, are also poor in essential amino acids; especially little in them lysine and tryptophan. Nevertheless, a purely vegetarian diet is not at all harmful, unless it consumes a slightly larger amount of vegetable proteins, sufficient to provide the body with essential amino acids. Most protein is found in plants in the seeds, especially in the seeds of wheat and various legumes. Young shoots, such as asparagus, are also rich in protein.Synthetic proteins in the diet. By adding small amounts of synthetic essential amino acids or proteins rich in them to incomplete proteins, such as corn proteins, one can significantly increase the nutritional value of the latter, i.e. thereby increasing the amount of protein consumed. Another possibility is to grow bacteria or yeasts on petroleum hydrocarbons with the addition of nitrates or ammonia as a source of nitrogen. The microbial protein obtained in this way can serve as feed for poultry or livestock, or can be directly consumed by humans. The third, widely used, method uses the physiology of ruminants. In ruminants, in the initial section of the stomach, the so-called. The rumen is inhabited by special forms of bacteria and protozoa that convert defective plant proteins into more complete microbial proteins, and these, in turn, after digestion and absorption turn into animal proteins. Urea, a cheap synthetic nitrogen-containing compound, can be added to livestock feed. Rumen-dwelling microorganisms use urea nitrogen to convert carbohydrates (of which there is much more in the feed) into protein. About a third of all nitrogen in livestock feed can come in the form of urea, which in essence means some chemical protein synthesis. In the USA, this method plays an important role as one of the ways to obtain protein.LITERATURE Murray R, Grenner D, Meyes P, Rodwell W. human biochemistry, tt. 12. M., 1993Alberts B., Bray D., Lewis J. et al. Molecular biology cells, tt. 13. M., 1994