Protein

INTRODUCTION TO,PROTEIN

Prepared byDr. Yogesh V. Ushir

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DEFINITION

Protein from the Greek proteios, meaning primary.

Proteins are the work horses of biological systems.–They play key roles in constructing and maintaining living cells

Our genes code for proteins

Proteins are polymers of amino acids

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SOURCES & QUALITY

Protein in food: different quantity & quality Nutritional value of protein: determined by

amino acid composition—Biological value (BV)

High biologicalvalueProtein that contains all essential amino acidsin sufficient proportions

Low biological valueProtein which is deficient (poor) in 1/moreessential amino acids

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ANIMAL & VEGETABLE PROTEIN

Animal proteinSources: 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 proteinSources: 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

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Animal protein = complete protein

Vegetable protein = incomplete protein

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MILK = HIGH QUALITY PROTEIN

Cow’s milk (and dairy products) = excellent source of high quality protein—All essential amino acids in proportions needed by the body

o Consists Whey protein (watery part of milk ) and casein (in milk salt of calcium )

Whey- liquid remaining after milk has been curdled and strained

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DIGESTION OF PROTEINS

Protein is hydrolysed (digested / cleaved) by particular protein-cleaving enzymes

Place of action: stomach & small intestine

Enzymes cleave protein in small units: —amino acids & short chain peptides

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ABSORPTION OF PROTEINS

Small units: absorbed by small finger-like projections (villi), lining the intestinal wall

Small peptides free amino acids

Free amino acids blood liver

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SYNTHESIS OF PROTEINS

Amino acids new (cell specific) protein = synthesis = continuing process

Body protein is continuously broken down (into amino acids) and re-synthesised (amino acids are re-built to protein)

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PROTEINS ARE:

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

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ROLES PLAYED BY PROTEINS INCLUDE

Enzymes (biologicalcatalysts) Hormones Storageproteins Transport proteins Structural proteins Protectiveproteins Contractile proteins Toxicproteins

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FUNCTION OF PROTEIN 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

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PROTEINS IN THE DIET

9 of the 20 amino acids must be obtained from the diet

These are referred to as the essential amino acids

Proteins are also the major source of nitrogen in the diet

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PROTEIN STRUCTURE

20 amino acids

Denatured

Quaternary

Tertiary

Secondary

Primary

Coded in DNA

Self assembly to a single (native) structure. Depends on primary structure and solution conditions

Common in foods. Many non-native forms depending on protein structure, solutionconditions (& history) and ingredient interactions

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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 oligo-

peptides

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AMINO ACIDS FOLDED IN A 3-DIMENSIONAL STRUCTURE

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

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• 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.

PROTEIN STRUCTURE AND FUNCTION

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THERE ARE 20 STANDARD AMINO

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PROPERTIES OF AMINO ACIDS

structure and chemical functionality

Chirality

acid-base properties

capacity to polymerize

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AMINO ACIDS: 2 GROUPS

Essential amino acids—Must be supplied with food—Cannot be synthesized (made) by our body

Non essential amino acids

—Can be synthesized by our body

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ESSENTIAL (INDISPENSABLE) AMINO ACIDS

Isoleucine Leucine Lysine Methionine Phenylalanine Threonine Tryptophan Valine Histidine

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AMINO ACIDS: 5 GROUPS

Aliphatic chains: Gly, Ala, Val, Leu, and Ile

hydrophobicity

Hydroxyl or sulfur side chains: Ser, Thr, Cys, Met

Aromatic: Phe, Trp, Tyr

Basic: His, Lys, Arg

Acidic and their amides: Asp, Asn, Glu, Gln

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ALIPHATIC (ALKANE) AMINO ACIDS

Proline(pro, P) –cyclic “iminoacid”

Glycine(gly, G)–only non-chiralamino acid, not hydrophobic

Alanine(ala, A) –R-group = methyl-group

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Valine(Val, V) –Think V!

Leucine(Leu, L) –

Isoleucine(Ile, I) -2 chiralcarbons

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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), H-

Bonding Tryptophan(Trp, W) –bicyclicindolering, H-

Bonding

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SULFUR CONTAINING AMINO ACIDS

Methionine (Met, M) –“start”amino acid, very hydrophobic

Cysteine (Cys,C) –sulfur in form of sulfhydroyl, important in disulfide linkages, weak acid, can form hydrogen bonds.

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ACIDIC AMINO ACIDS

Contain carboxyl groups (weaker acids than α-carboxyl-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 –

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BASIC AMINO ACIDS

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

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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

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NONSTANDARD AMINO ACIDS

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These include the amino acid derivatives found in proteins;

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

argininesynthesis

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FUNCTIONAL SIGNIFICANCE

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

nucleophilesduring enzymatic catalysis–Disulfide bonds

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STEREOCHEMISTRY OF AMINO ACIDS

Amino acids are chiral (asymmetric)–Presence of an

asymmetric carbon atom

Natural proteins are made only from left-handed amino acids

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PEPTIDE BOND FORMATION0

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USE OF SYMBOLS IN REPRESENTING A PEPTIDES

In peptides the free –NH3+ is on the left while

free –COO – is on the right

Tripeptide –NH3

+ - glutamate-cystein-glycine-COO – - E - C – G one letter

symbols Glu- Cys – Gly three letter

symbols Glutamyl-cysteinyl-glycine peptide name

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PROPERTIES OF AMINO ACIDS PHYSICAL

Solubility Melting point Optical rotation Taste Acid base (ampholytes)

CHEMICAL Reaction due to –COOH grs.

Decarboxylation With ammonia

Reaction due to –NH2 grps. With ninhydrin Color reaction Transamination Oxidative deamination

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ACID-BASE PROPERTIES OF AMINO ACIDS

Amino acids are zwitterions:

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TITRATION OF AMINO ACIDS: ACID-BASE TITRATION

pK1: carboxylic acid = 2.4

pK2: amino group = 9.6

pI= (pK1+ pK2)/2

So, PI for leucine=6

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ABSORPTION SPECTRA OF AROMATIC AMINO ACIDS

Trp → λmax of 278nmTyr → λmax of 275nmPhe → λmax of 260nm

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DISULFIDE BRIDGE

- CH2 – SH + - CH2 – SH

- 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. Although Cys is a polar AA, the di-sulfide linked

residues (Cystine) are strongly hydrophobic.

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NINHYDRIN REACTION

– More common for amino acids

- Purple, blue or violet derivatives, 570nm

– Yellow for proline, 440 nm

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BONDS RELATING TO PROTEIN STRUCTURES

1. Strong bonds 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

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BIOLOGICALLY ACTIVE PEPTIDES 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 nature

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PROTEINS

May be defined as the high molecular weight mixed polymer s of alpha-amino acids joined together with peptide linkage

-(CO – NH) –- The term protein is generally used for a

polypeptide containing more than (50 ?) 100 amino acids.

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CLASSIFICATION1. SIMPLE PROTEINS

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

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2. CONJUGATED PROTEINS a. Nucleoproteins:

nucleic acid (DNA & RNA) is the prosthetic group

E.g.; nucleohistones, nucleoprotamines

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)

c. lipoproteins: the prosthetic group is lipid E.g.; serum lipoproteins, membrane LP

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2. CONJUGATED PROTEINS (CONTINUE…)

d. Phosphoproteins: phosphoric acid is the prosthetic group

E.g.; casein, viteline (egg yolk)

e. chromoproteins: prosthetic group is colored in nature

E.g.; Hb, cytochromes

f. metalloproteins: these protein contains metal ions, Fe, Zn, Cu, Mg

E.g.; ceruloproteins (Cu), carbonic anhydrase (Zn)

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3. DERIVED PROTEINS

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

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NUTRITONAL CLASSIFICATION

Complete proteins

Incomplete proteins

Already discuss earlier

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PROTEIN HYDROLYZING ENZYMES

Pepsin- in gastric juice

Trypsin-, chymotrypsin and carboxypeptidases- in pancreatic juice

Amino-peptidases, dipeptidases and polypeptidases-in intestinal juice

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STRUCTURE OF PROTEINS Primary- is the linear sequence of amino acids

forming the backbone of proteins (polypeptides)

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

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TESTS FOR PROTEINS

a. Color reactions Biuret test Ninhydrin test

b. co-agulation reactions Heat coagulation

c. Ppt. reactions Gives gelatinous ppt with solid ammonium

sulphate

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ESTIMATION OF PROTEINS

Quantitative estimation of proteins of foods and other biological materials are performed by the following methods-

Kjeldhal method

Colorimetric method

Electrophoretic method

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Thank u… Thank u… meet u again meet u again in synthesisin synthesis

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