Protein

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Protein

  1. 1. 09/07/2012 INTRODUCTION TO, PROTEIN H.N.S.I.P.E.R., Rajkot 1 Prepared by Dr. Yogesh V. Ushir
  2. 2. DEFINITION Proteins are the work horses of biological systems. H.N.S.I.P.E.R., Rajkot Protein from the Greek proteios, meaning primary.  09/07/2012  –They play key roles in constructing and maintaining living cells  Our genes code for proteins  Proteins are polymers of amino acids 2
  3. 3. SOURCES & QUALITY 09/07/2012  Protein biologicalvalue Protein that contains all essential amino acids in sufficient proportions  Low biological value Protein which is deficient (poor) in 1/more essential amino acids H.N.S.I.P.E.R., Rajkot in food: different quantity & quality  Nutritional value of protein: determined by amino acid composition —Biological value (BV)  High 3
  4. 4. ANIMAL & VEGETABLE PROTEIN Animal protein Sources: e.g. meat, poultry, fish, eggs, milk, dairy products — Almost same proportion of each essential amino acid as human protein —Complete protein: high BV  Vegetable protein Sources: e.g. vegetables, legumes, plants, grains, nuts, seeds — Deficient in 1/more essential amino acids —Incomplete protein: low BV — Deficient (poor) amino acid = limiting amino acid 09/07/2012  H.N.S.I.P.E.R., Rajkot 4
  5. 5. Animal protein = complete protein  Vegetable protein = incomplete protein 09/07/2012  H.N.S.I.P.E.R., Rajkot 5
  6. 6. MILK = HIGH QUALITY PROTEIN Consists Whey protein (watery part of milk ) and casein (in milk salt of calcium )  Whey- liquid remaining after milk has been curdled and strained H.N.S.I.P.E.R., Rajkot Cow’s milk (and dairy products) = excellent source of high quality protein —All essential amino acids in proportions needed by the body o 09/07/2012  6
  7. 7. DIGESTION OF PROTEINS Place of action: stomach & small intestine  Enzymes cleave protein in small units: —amino acids & short chain peptides H.N.S.I.P.E.R., Rajkot Protein is hydrolysed (digested / cleaved) by particular protein-cleaving enzymes  09/07/2012  7
  8. 8. ABSORPTION OF PROTEINS Small peptides  Free amino acids H.N.S.I.P.E.R., Rajkot Small units: absorbed by small finger-like projections (villi), lining the intestinal wall  09/07/2012  free amino acids blood liver 8
  9. 9. SYNTHESIS OF PROTEINS Body protein is continuously broken down (into amino acids) and re-synthesised (amino acids are re-built to protein) H.N.S.I.P.E.R., Rajkot Amino acids new (cell specific) protein = synthesis = continuing process  09/07/2012  9
  10. 10. PROTEINS ARE: 09/07/2012 H.N.S.I.P.E.R., Rajkot Polypeptides + (cofactors, coenzymes, prosthetic groups, other modifications in animal)  Polypeptides are covalently linked a-amino acids  Cofactors are non-amino acid components e.g. metal ions like Zn2+in carboxypeptidase  Coenzymes are organic cofactors e.g. nucleotides in lactate dehydrogenase  Prosthetic groups are covalently attached cofactors e.g. hemein myoglobin  10
  11. 11. ROLES PLAYED BY PROTEINS INCLUDE 09/07/2012 H.N.S.I.P.E.R., Rajkot Enzymes (biologicalcatalysts)  Hormones  Storageproteins  Transport proteins  Structural proteins  Protectiveproteins  Contractile proteins  Toxicproteins  11
  12. 12. FUNCTION OF PROTEIN    H.N.S.I.P.E.R., Rajkot  Major functional & structural component of all cells in body required for growth, maintenance, renewal (repair) of all body cells Functions as enzyme, transport carrier, hormone —All enzymes, many hormones, blood transport molecules, hair, fingernails, etc. are proteins Amino acids play a role as precursors (forerunner) for e.g. many enzymes, hormones, vitamins Constituent of antibodies which protect against infection & disease Supplies energy in particular circumstances (e.g. in malnourishment), —When fat & carbohydrate intake is inadequate 12 09/07/2012 
  13. 13. PROTEINS IN THE DIET  These are referred to as the essential amino acids  Proteins are also the major source of nitrogen in the diet H.N.S.I.P.E.R., Rajkot 9 of the 20 amino acids must be obtained from the diet 09/07/2012  13
  14. 14.   PROTEIN STRUCTURE Coded in DNA Secondary Self assembly to a single (native) structure. Depends on primary Tertiary structure and solution conditions Quaternary Denatured Common in foods. Many non-native forms depending on protein structure, solution conditions (& history) and ingredient interactions H.N.S.I.P.E.R., Rajkot Primary 09/07/2012 20 amino acids 14
  15. 15. Protein consists of amino acids Amino acid (AA)= The basic organic compounds of proteins that are linked by peptide bonds (make up true protein). Can be compared with a chain of beads ---------etc  poly-peptide = protein    Building unit =amino acid   A few amino acids linked together -------di-, tri-, or oligopeptides H.N.S.I.P.E.R., Rajkot  09/07/2012  15
  16. 16. AMINO ACIDS FOLDED IN A 3DIMENSIONAL STRUCTURE 09/07/2012 H.N.S.I.P.E.R., Rajkot Protein constituted by 5 elements in proportion;  C = carbon - 50-55%  H = hydrogen - 6-7%  O = oxygen - 19-24%  S = sulfur -0-4%  N = nitrogen - 13-19% N= key element for building tissues 16
  17. 17. PROTEIN STRUCTURE AND FUNCTION 09/07/2012 H.N.S.I.P.E.R., Rajkot • Protein are polymers of a-amino acids • The amino acids used to make proteins are 2-amino-carboxylic acids •The α- (alpha) carbon is the carbon to which a functional group is attached. 17
  18. 18. ACIDS H.N.S.I.P.E.R., Rajkot 20 STANDARD AMINO 09/07/2012 THERE ARE 18
  19. 19. PROPERTIES OF AMINO ACIDS and chemical functionality  acid-base  capacity properties H.N.S.I.P.E.R., Rajkot  Chirality 09/07/2012  structure to polymerize 19
  20. 20. AMINO ACIDS: 2 GROUPS    Non essential amino acids —Can be synthesized by our body H.N.S.I.P.E.R., Rajkot Essential amino acids —Must be supplied with food —Cannot be synthesized (made) by our body 09/07/2012  20
  21. 21. ESSENTIAL (INDISPENSABLE) AMINO ACIDS 09/07/2012 H.N.S.I.P.E.R., Rajkot Isoleucine  Leucine  Lysine  Methionine  Phenylalanine  Threonine  Tryptophan  Valine  Histidine  21
  22. 22. AMINO ACIDS: 5 GROUPS Aliphatic chains: hydrophobicity  Hydroxyl or sulfur side chains: Ser, Thr, Cys, Met  Aromatic: H.N.S.I.P.E.R., Rajkot Gly, Ala, Val, Leu, and Ile 09/07/2012  Phe, Trp, Tyr  Basic: His, Lys, Arg  Acidic and their amides: Asp, Asn, Glu, Gln 22
  23. 23. ALIPHATIC (ALKANE) AMINO ACIDS  Glycine(gly, G)–only non-chiralamino acid, not hydrophobic  Alanine(ala, A) –R-group = methyl-group H.N.S.I.P.E.R., Rajkot Proline(pro, P) –cyclic “iminoacid” 09/07/2012  23
  24. 24. Valine(Val, V) –Think V!  Leucine(Leu, L) –  Isoleucine(Ile, I) -2 chiralcarbons 09/07/2012  H.N.S.I.P.E.R., Rajkot 24
  25. 25. AROMATIC AMINO ACIDS All very hydophobic  All contain aromatic group  Absorb UV at 280 nm  Phenylalanine(Phe, F)  Tyrosine(Tyr,Y) –-OH ionizable(pKa= 10.5), HBonding  Tryptophan(Trp, W) –bicyclicindolering, HBonding  09/07/2012 H.N.S.I.P.E.R., Rajkot 25
  26. 26. SULFUR CONTAINING AMINO ACIDS  Cysteine (Cys,C) –sulfur in form of sulfhydroyl, important in disulfide linkages, weak acid, can form hydrogen bonds. H.N.S.I.P.E.R., Rajkot Methionine (Met, M) –“start”amino acid, very hydrophobic 09/07/2012  26
  27. 27. ACIDIC AMINO ACIDS H.N.S.I.P.E.R., Rajkot  09/07/2012 Contain carboxyl groups (weaker acids than αcarboxyl-group)  Negatively charged at physiological pH, present as conjugate bases (therefore –ate not –ic acids)  Carboxyl groups function as nucleophiles in some enzymatic reactions  Aspartate–  Glutamate – 27
  28. 28. BASIC AMINO ACIDS 09/07/2012 H.N.S.I.P.E.R., Rajkot Hydrophillic nitrogenous bases  Positively charged at physiological pH  Histidine–imidazole ring protonated/ionized, only amino acid that functions as buffer in physio range.  Lysine –di-amino acid, protonated at pH 7.0  Arginine-guianidinium ion always protonated, most basic amino acid  28
  29. 29. POLAR UNCHARGED AMINO ACIDS Serine(Ser, S) –looks like Ala w/ -OH  Threonine(Thr, T) –2 chiralcarbons  Asparagine(Asn, N) –amide of aspartic acid  Glutamine (Gln, Q) –amide of glutamicacid 09/07/2012  H.N.S.I.P.E.R., Rajkot 29
  30. 30. H.N.S.I.P.E.R., Rajkot ? 09/07/2012 NONSTANDARD AMINO ACIDS 30
  31. 31. These include the amino acid derivatives found in proteins; 09/07/2012 H.N.S.I.P.E.R., Rajkot Hydroxylysine, hydroxyproline -collagen  Carboxyglutamate -blood-clotting proteins  Pyroglutamate -bacteriorhodopsin  Phosphorylatedamino acids -signaling device  D-alanine, D-glutamic acid - bacterial cell wall polypeptides  Gamaaminobutyricacid (GABA)- neurotransmitter  Homoserine -intermediate in amino acid metabolism  Ornithine - intermediate in 31 argininesynthesis 
  32. 32. FUNCTIONAL SIGNIFICANCE 09/07/2012 H.N.S.I.P.E.R., Rajkot Hydrophobic amino acids: encountered in the interior of proteins shielded from direct contact with water  Hydrophilic amino acids: generally found on the exterior of proteins as well as in the active centers of enzymes  Imidazole group: act as either proton donor or acceptor at physiological pH  –Reactive centers of enzymes  Primary alcohol and thiol groups: act as nucleophiles during enzymatic catalysis –Disulfide bonds 32
  33. 33. STEREOCHEMISTRY OF AMINO ACIDS –Presence of an asymmetric carbon atom  H.N.S.I.P.E.R., Rajkot Amino acids are chiral (asymmetric) 09/07/2012  Natural proteins are made only from lefthanded amino acids 33
  34. 34. PEPTIDE BOND FORMATION 09/07/2012 H.N.S.I.P.E.R., Rajkot 34
  35. 35. USE OF SYMBOLS IN REPRESENTING A PEPTIDES  Tripeptide  –NH3+ E - glutamate-cystein-glycine-COO – - -C–G  Glu- Cys – Gly  Glutamyl-cysteinyl-glycine one letter symbols three letter symbols peptide name H.N.S.I.P.E.R., Rajkot In peptides the free –NH3+ is on the left while free –COO – is on the right 09/07/2012  35
  36. 36. PROPERTIES OF AMINO ACIDS  PHYSICAL  Melting  CHEMICAL  Reaction due to –COOH grs.  Decarboxylation  With ammonia  Reaction due to –NH2 grps.  With ninhydrin  Color reaction  Transamination  Oxidative deamination H.N.S.I.P.E.R., Rajkot point  Optical rotation  Taste  Acid base (ampholytes) 09/07/2012  Solubility 36
  37. 37. ACID-BASE PROPERTIES OF AMINO ACIDS Amino acids are zwitterions: 09/07/2012  H.N.S.I.P.E.R., Rajkot 37
  38. 38. TITRATION OF AMINO ACIDS: ACID-BASE TITRATION 09/07/2012 H.N.S.I.P.E.R., Rajkot pK1: carboxylic acid = 2.4 pK2: amino group = 9.6 pI= (pK1+ pK2)/2 So, PI for leucine=6 38
  39. 39. ABSORPTION SPECTRA OF AROMATIC AMINO ACIDS 09/07/2012 H.N.S.I.P.E.R., Rajkot Trp → λmax of 278nm Tyr → λmax of 275nm Phe → λmax of 260nm 39
  40. 40. DISULFIDE BRIDGE CH2 – SH + - CH2 – SH [O] H.N.S.I.P.E.R., Rajkot - 09/07/2012 - - CH2 – S– S - CH2    Two cysteine molecules under oxidizing conditions form the Disulfide bridge Disulfide bonds between Cys residues stabilize the structures of many proteins. 40 Although Cys is a polar AA, the di-sulfide linked residues (Cystine) are strongly hydrophobic.
  41. 41. NINHYDRIN REACTION  - Purple, blue or violet derivatives, 570nm  – Yellow for proline, 440 nm H.N.S.I.P.E.R., Rajkot – More common for amino acids 09/07/2012  41
  42. 42. BONDS RELATING TO PROTEIN STRUCTURES  Peptide bond Disulphide bond 2. Weak bonds   Hydrogen bonds- H-atoms between nirogen & carbony oxygen Hydrophobic bonds- non-polar side chains of neutral amino acids are closely associated with one another in proteins H.N.S.I.P.E.R., Rajkot  09/07/2012 1. Strong bonds 42
  43. 43. BIOLOGICALLY ACTIVE PEPTIDES 09/07/2012 Glutathione - 3peptides  Thyrotropin releasin harmone - 3peptides  Oxytocin - 9 amino acids  Vasopressin (ADH) - 9 amino acids  Angiotensins - 10 amino acids  Methionine enkephalin - 5peptides  Bradykinin - 9 amino acids  Kallidin - 10 amino acids  Aspatame - dipeptide  Peptide antiibiotic- antibiotic such as gramicidine, bacitracin, tyrocidin & actinomycin are peptide in 43 nature  H.N.S.I.P.E.R., Rajkot
  44. 44. PROTEINS - (CO – NH) – H.N.S.I.P.E.R., Rajkot May be defined as the high molecular weight mixed polymer s of alpha-amino acids joined together with peptide linkage 09/07/2012  The term protein is generally used for a polypeptide containing more than (50 ?) 100 amino acids. 44
  45. 45. CLASSIFICATION 1. SIMPLE PROTEINS 09/07/2012 H.N.S.I.P.E.R., Rajkot a. Globular proteins: these are spherical or oval shape, soluble in water or other solvents and digestible  E.g.; albumin, globulin, glutelin, prolamines, histones, globins, protamines b. Fibrous proteins: these are fibre like in shape, insoluble in water & resistance to digetion.  E.g.; collagens, elastins, keratins 45
  46. 46. 2. CONJUGATED PROTEINS c. lipoproteins: the prosthetic group is lipid  E.g.; serum lipoproteins, membrane LP H.N.S.I.P.E.R., Rajkot b. Glycoproteins: the prosthetic group is carbohydrates, which is less than 4 % of proteins. The term mucoproteins is used if > 4% of carbohydrates  E.g.; mucin, ovomucoid (egg white) 09/07/2012 a. Nucleoproteins: nucleic acid (DNA & RNA) is the prosthetic group  E.g.; nucleohistones, nucleoprotamines 46
  47. 47. 2. CONJUGATED PROTEINS (CONTINUE…) f. metalloproteins: these protein contains metal ions, Fe, Zn, Cu, Mg  E.g.; ceruloproteins (Cu), carbonic anhydrase (Zn) H.N.S.I.P.E.R., Rajkot e. chromoproteins: prosthetic group is colored in nature  E.g.; Hb, cytochromes 09/07/2012 d. Phosphoproteins: phosphoric acid is the prosthetic group  E.g.; casein, viteline (egg yolk) 47
  48. 48. 3. DERIVED PROTEINS 09/07/2012 H.N.S.I.P.E.R., Rajkot a. Primary derived proteins: are the denatured or coagulated or first hydrolyzed product of proteins  E.g.; proteans (fibrin, fibrinogen), coagulated protein (cooked protein), metaprotein b. Secondary derived proteins: are the progressive hydrolytic product of protein hydrolysis  E.g.; proteoses, peptones, polypeptides & peptides 48
  49. 49. NUTRITONAL CLASSIFICATION proteins proteins H.N.S.I.P.E.R., Rajkot Incomplete 09/07/2012 Complete Already discuss earlier 49
  50. 50. PROTEIN HYDROLYZING ENZYMES  Trypsin-, chymotrypsin and carboxypeptidasesin pancreatic juice  Amino-peptidases, dipeptidases and polypeptidases-in intestinal juice H.N.S.I.P.E.R., Rajkot Pepsin- in gastric juice 09/07/2012  50
  51. 51. STRUCTURE OF PROTEINS  Secondary- the spatial arrangement of proteins by twisting of the polypeptide chain  Tertiary- The three di-mentional structure of a protein  Quaternary- some of the proteins are composed of two or more polypeptides chains refereed to as subunits. The spatial arrangement of these subunits is known as Quaternary proteins H.N.S.I.P.E.R., Rajkot Primary- is the linear sequence of amino acids forming the backbone of proteins (polypeptides) 09/07/2012  51
  52. 52. TESTS FOR PROTEINS b. co-agulation reactions  Heat coagulation H.N.S.I.P.E.R., Rajkot Biuret test  Ninhydrin test  09/07/2012 a. Color reactions c. Ppt. reactions  Gives gelatinous ppt with solid ammonium sulphate 52
  53. 53. ESTIMATION OF PROTEINS Colorimetric method  H.N.S.I.P.E.R., Rajkot  09/07/2012 Quantitative estimation of proteins of foods and other biological materials are performed by the following methods Kjeldhal method Electrophoretic method 53
  54. 54. 09/07/2012 Thank u… meet u again in synthesis H.N.S.I.P.E.R., Rajkot 54

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