The source of protein structures is the Protein Data Bank. The unit of classification of structure in SCOP is the protein domain.

Transcription

1 UNIT 14 PROTEINS

2 DEFINITION A large molecule composed of one or more chains of amino acids in a specific order; the order is determined by the base sequence of nucleotides in the gene that codes for the protein. Proteins are required for the structure, function, and regulation of the body's cells, tissues, and organs; and each protein has unique functions. Examples are hormones, enzymes, and antibodies

3 CLASSIFICATION OF PROTEINS The source of protein structures is the Protein Data Bank. The unit of classification of structure in SCOP is the protein domain. The shapes of domains are called "folds" in SCOP. Domains belonging to the same fold have the same major secondary structures in the same arrangement with the same topological connections. Short descriptions of each fold are given. For example, the "Globin-like" fold is described as core: 6 helices; folded leaf, partly opened.

4 CLASSES: a. All alpha proteins domains consisting of α-helices b. All beta proteins domains consisting of ß-sheets c. Alpha and beta proteins. Mainly parallel beta sheets (beta-alpha-beta units) d. Alpha and beta proteins. Mainly antiparallel beta sheets (segregated alpha and beta regions) e. Multi-domain proteins (alpha and beta). Folds consisting of two or more domains belonging to different classes f. membrane and cell surface proteins and peptides. Does not include proteins in the immune system g. Small proteins

5 KINDS OF PROTEINS CLASSIFICATION BASED UPON FUNCTION Enzymes (catalytic proteins) Lactase, ribonuclease, pyruvic dehydrogenase, fumarase, proteinase Structural proteins Collagen, elastin, keratin Regulatory or hormonal proteins Insulin, adrenaline Transport proteins Hemoglobin, myoglobin Genetic proteins Nucleoproteins, histones Immune Proteins Gamma Globulin, IG's, (Ab's) Contractile Proteins Actin, myosin Storage Proteins Zein, ovalbumin, casein

6 CLASSIFICATION BASED UPON SHAPE Fibrous Proteins - Long thread-like molecules whose helical strands often form fibers or sheets; E.g. Collagen, elastin, keratin Globular Proteins - Generally soluble in aqueous media; spheroid or ovoid shape; further classified on the basis of solubility - water soluble, heat coagulable, include proteins in blood serum, egg white, milk, soluble in dilute salt solutions.

7 CLASSIFICATION - PHYSICOCHEMICAL PROPERTIES: Simple Proteins - Yield only amino acids on hydrolysis Albumins, Globulins, Glutelins Soluble in dilute acids and alkalies Scleroproteins - Nonsoluble proteins such as Collagen Prolamines Soluble in alcohol, insoluble in water; found in seeds of plants Histones Soluble in water, dilute acids, and alkalies; contain a large portion of basic amino acids such as globin of hemoglobin Protamines Basic proteins which are essentially large polypeptides

8 PROTEIN ELECTROPHORESIS Protein Electrophoresis is a method in which a mixture of proteins can be separated and analyzed. Electrophoresis is based on the mobility of ions in an electric field. The charge distribution of the molecules is critical in the separation of all electrophoresis. In an electric field, electrophoresis is a passage of charged molecules in solution. Positively charged ions have tendency toward a negative electrode and inversely, negatively charged ions have tendency toward a positive electrode. The molecular weight results to a molecular friction which is directly proportional to the molecular charge and its voltage and inversely proportional to a molecule s mobility in an electric field.

9 PEPTIDE BOND: Two amino acid molecules can be covalently joined through a substituted amide linkage, termed a peptide bond, to yield a dipeptide. Process can then continue to join other amino acids and yield in an amino acid chain. When there are few amino acids in a chain, it is called an oligopeptide, when there are many it is called a polypeptide. An amino acid unit in a peptide is often called a residue.

10 Proteins are polymers of amino acids. Proteins differ from each other according to the type, number and sequence of amino acids that make up the polypeptide backbone. As a result they have different molecular structures, nutritional attributes and physiochemical properties. They are a major source of energy, as well as containing essential amino-acids, such as lysine, tryptophan, methionine, leucine, isoleucine and valine, which are essential to human health, but which the body cannot synthesize. Proteins are also the major structural components of many natural foods, often determining their overall texture, e.g., tenderness of meat or fish products

11 ISOLATED PROTEINS: Isolated proteins are often used in foods as ingredients because of their unique functional properties, i.e., their ability to provide desirable appearance, texture or stability. Typically, proteins are used as gelling agents, emulsifiers, foaming agents and thickeners. Many food proteins are enzymes which are capable of enhancing the rate of certain biochemical reactions. These reactions can have either a favorable or detrimental effect on the overall properties of foods. Food analysts are interested in knowing the total concentration, type, molecular structure and functional properties of the proteins in foods.

12 Kjeldahl method The amount of protein present is then calculated from the nitrogen concentration of the food. The same basic approach is still used today, although a number of improvements have been made to speed up the process and to obtain more accurate measurements. It is usually considered to be the standard method of determining protein concentration. The Kjeldahl method can conveniently be divided into three steps: digestion, neutralization and titration

13 NEUTRALIZATION After the digestion has been completed the digestion flask is connected to a recieving flask by a tube. The solution in the digestion flask is then made alkaline by addition of sodium hydroxide, which converts the ammonium sulfate into ammonia gas: The ammonia gas that is formed is liberated from the solution and moves out of the digestion flask and into the receiving flask - which contains an excess of boric acid. The low ph of the solution in the receiving flask converts the ammonia gas into the ammonium ion, and simultaneously converts the boric acid to the borate ion: NH3 + H3BO3 (boric acid) NH4+ + H2BO3- (borate ion) (3)

14 ADVANTAGES AND DISADVANTAGES Advantages. The Kjeldahl method is widely used internationally and is still the standard method for comparison against all other methods. Its universality, high precision and good reproducibility have made it the major method for the estimation of protein in foods. Disadvantages. It does not give a measure of the true protein, since all nitrogen in foods is not in the form of protein. Different proteins need different correction factors because they have different amino acid sequences. The use of concentrated sulfuric acid at high temperatures poses a considerable hazard, as does the use of some of the possible catalysts The technique is time consuming to carry-out.

15 ISOELECTRIC POINT The isoelectric point of a protein is the ph where the net charge on the protein is zero. Proteins tend to aggregate and precipitate because there is no electrostatic repulsion keeping them apart. Proteins have different isoelectric points because of their different amino acid sequences (i.e., relative numbers of anionic and cationic groups), and thus they can be separated by adjusting the ph of a solution. When the ph is adjusted to the pi of a particular protein it precipitates leaving the other proteins in solution.

16 Solvent Fractionation The solubility of a protein depends on the dielectric constant of the solution that surrounds it because this alters the magnitude of the electrostatic interactions between charged groups. The amount of organic solvent required to cause precipitation depends on the protein and therefore proteins can be separated on this basis. The optimum quantity of organic solvent required to precipitate a protein varies from about 5 to 60%. Solvent fractionation is usually performed at 0oC or below to prevent protein are mixed with water.