Proteins

unit vi

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 Proteins

• Proteins are nitrogenous “macromolecules” composed of many amino

acids.

• The term protein is derived from Greek word “Proteios” , which means
“primary”, or “holding first place”.

• They are named so because proteins are the most important of biological
substances and are fundamental structural components of the body.

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 Composition of Proteins
• In addition to C, H, and O, proteins also contain N. The nitrogen content is around 16% of
the molecular weight of proteins.

• Small amounts of S and P are also present. Few proteins contain other elements such as
I, Cu, Mn, Zn and Fe, etc.

• Protein molecules are very large molecules made up of small units called Amino acids.

• More than 300 hundred amino acids have been described but only 20 amino acids have
been found to be present in mammalian tissues and take part in protein synthesis.

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 Amino acids

R is called a side chain and can be a hydrogen, aliphatic, aromatic or

heterocyclic group. Each amino acid has an amino group –NH2, a carboxylic
acid group -COOH and a hydrogen atom each attached to carbon
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 Amino Acids- Classification and Structure
• Amino acids can be classified into 3 groups depending on their reaction in solution.

• A. Neutral

• B. Acidic

• C. Basic

• A. Neutral amino acids:

• This is the largest group of amino acids and can be further subdivided into aliphatic,
aromatic, heterocyclic and S-containing amino acids.

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 (a) Aliphatic Amino Acids

• Glycine (Gly: G) or α-amino acetic acid.

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 (a) Aliphatic Amino Acids

• Alanine (Ala: A) or α-amino propionic acid.

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 (a) Aliphatic Amino Acids

• Valine (Val: V) or α-amino-isovaleric acid.

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 (a) Aliphatic Amino Acids

• Leucine (Leu: L) or α-amino-isocaproic acid.

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 (a) Aliphatic Amino Acids

• Isoleucine (Ile: I) or α-amino-β-methyl valeric acid.

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 (a) Aliphatic Amino Acids
• Serine (Ser: S) or α-amino-β-hydroxy propionic acid.

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 (a) Aliphatic Amino Acids
• Threonine (Thr: T) or α-amino-β-hydroxybutyric acid.

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 (b) Aromatic Amino Acids
• Second subgroup of neutral amino acids consists of aromatic amino
acids.
• Phenylalanine (Phe: F) or α-amino-β-phenyl propionic acid.

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 (b) Aromatic Amino Acids
• Tyrosine (Tyr: Y) or α-amino-β-parahydroxy phenylpropionic acid.

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 (c) Heterocyclic Amino Acids
• Third group belongs to heterocyclic amino acids.
• Tryptophan (Trp: W) or α-amino-β-3-indole propionic acid.
• This amino acid is often considered as aromatic amino acid since it
has aromatic ring in its structure.

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 (c) Heterocyclic Amino Acids
• Histidine (His: H) or α-amino-β-imidazole propionic acid.
• Histidine is basic in solution on account of the imidazole ring and
often considered as Basic Amino acid.

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 (d) Imino Acids
• Proline (Pro: P) or Pyrrolidone-2-carboxylic acid.

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 (d) Imino Acids
• Hydroxyproline (Hyp) or 4 hydroxy pyrrolidone-2 carboxylic acid.

• Proline and Hydroxyproline do not have a free –NH2 group but only a basic pyrrolidone ring in
which the Nitrogen of the Imino group is in a ring but can still function in the formation of
peptides. These amino acids are therefore called as imino acids.

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 (e) ‘S’ containing amino acids
• The fourth subgroup of neutral amino acids contains two sulphur
containing amino acids.
• Cysteine (Cys: C) or α-amino-β-mercaptopropionic acid.

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 (e) ‘S’ containing amino acids
• Two molecules of cysteine make cystine (cys-cys). The S–S linkage is
called as disulphide bridge.

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 (e) ‘S’ containing amino acids
• Methionine (Met: M) or α-amino γ-methylthio-η-butyric acid.

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 (B) Acidic amino acids
• These amino acids have two -COOH groups and one – NH2 group.
They are therefore mono amino di carboxylic acids.
• Aspartic acid (Asp: D) or α-amino succinic acid.

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 (B) Acidic amino acids
• Glutamic Acid (Glu: E) or α-aminoglutaric acid. First isolated from
wheat protein glutein.

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 (C) Basic amino acids

• This class of amino acids consists of those amino acids which have
one – COOH group and two –NH2 groups. Thus they are di amino
mono carboxylic acids. Arginine, lysine and hydroxylysine are included
in this group.

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 (C)Basic amino acids
• Arginine (Arg: R) or α-amino- δ-guanidino-n-valeric acid.

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 (C)Basic amino acids
• Lysine (Lys: K) or α-ε-diamino caproic acid.

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 (C)Basic amino acids
• Hydroxylysine (Hyl) or α-ε-diamino-δ-hydroxy-n-valeric acid.

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 Non-standard Amino Acids
• The compounds similar to basic structure of amino acids but do not occur in
proteins. Examples are:
• Ornithine and citrulline: They are intermediates in urea cycle
• Thyroxine (T4) and Tri-iodo Thyronine (T3): Thyroid hormones synthesised from
tyrosine.
• Gamma-aminobutyric acid (GABA): A neurotransmitter produced from glutamic
acid.
• S-adenosyl methionine (SAM): These are methyl donor formed from L-
methionine
• 3, 4-dihydroxy phenyl alanine (DOPA): A precursor of melanine pigment.
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 Functions of Amino Acids
• Apart from making proteins and peptides, amino acids serve variety of functions.
• Some amino acids are converted to carbohydrates and are called as glucogenic
amino acids.
• Specific amino acids give rise to specialised products, e.g. Tyrosine forms
hormones such as thyroid hormones,(T3, T4), epinephrine and norepinephrine
and a pigment called melanin.
• Tryptophan can synthesise a vitamin called niacin.
• Glycine and cysteine help in synthesis of Bile salts.
• Glutamate, cysteine and glycine synthesise glutathione.

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 Functions of Amino Acids
• Histidine changes to histamine on decarboxylation.
• Serotonin is formed from tryptophan.
• Pyrimidines and purines use several amino acids for their synthesis
such as aspartate and glutamine for pyrimidines and glycine, aspartic
acid, Glutamine and serine for purine synthesis.
• Methionine acts as “active” methionine (S-adenosyl-methionine) and
transfers methyl group to various substances by transmethylation.
• (e) Cystine and methionine are sources of sulphur.

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 Nutritional classification (Amino Acids)
• Nutritionally, amino acids are of three types:
• (a) Essential
• (b) Non-essential
• (c) Semi-essential amino acids
• (a) Essential amino acids: These are the ones which are not synthesized by the
body and must be taken in diet. They include valine, leucine, isoleucine,
phenylalanine, threonine, tryptophan, methionine and lysine. For remembering
the following formula is used—MATT VIL PHLY.
• (b) Non-essential amino acids: They can be synthesized by the body and may not
be the requisite components of the diet.
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 Semi-essential amino acids

• (c) Semi-essential amino acids: These are growth promoting factors

since they are not synthesized in sufficient quantity during growth.

• They include arginine and histidine. They become essential in

growing children, pregnancy and lactating women.

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 Amphoteric Nature of Amino acids
• The -NH2 and -COOH groups of amino acids are ionizable groups.
• Depending on the pH of the solution these groups act as proton
donors (acids) or proton acceptors (bases).
• Substances having this dual(acid-base) nature are called amphoteric
and therefore amino acids are called as ampholytes (amphoteric
electrolytes).
• At a specific pH the amino acid carries both the charges in equal
number and exists as dipolar ion or “Zwitterion” (German for “hybrid
ion”).
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 Zwitterion

Nonionic and zwitterionic forms of amino acids. The non-ionic form does not occur in significant
amounts in aqueous solutions. The zwitterion predominates at neutral pH. A zwitterion can act as
either an acid (proton donor) or a base (proton acceptor).
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 Isoelectric pH

• The pH at which the net charge on an amino acid is zero, i.e. positive
charges and negative charges on the protein/amino acid molecule
equalizes, is called pI or isoelectric pH.

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 Classification of Proteins

• Proteins are classified:

1. On the basis of shape and size

2. On the basis of functional properties

3. On the basis of solubility and physical properties

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 1. On the basis of shape and size

a) Fibrous proteins

• When the axial ratio of length: width of a protein molecule is more than 10, it is
called a fibrous protein. Examples: α-keratin from hair, collagen.

b) Globular protein

• When the axial ratio of length: width of a protein molecule is less than 10, it is
called as globular protein. Examples: Myoglobin, haemoglobin, ribonuclease, etc.

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 2. On the basis of functional properties:
a) Defense proteins: Immunoglobulins involved in defence mechanisms.

b) Contractile proteins: Proteins of skeletal muscle involved in muscle contraction and

relaxation.

c) Respiratory proteins: Involved in the function of respiration, like haemoglobin, myoglobin,

cytochromes.

d) Structural proteins: Proteins of skin, cartilage, nail.

e) Enzymes: Proteins acting as enzymes.

f) Hormones: Proteins acting as hormones.

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3. On the basis of solubility and physical
properties
• A. Simple proteins: These are proteins which on complete hydrolysis yield
only amino acids.

• B. Conjugated proteins: These are proteins which in addition to amino acids

in their structure, contain a non-protein group called prosthetic group

• C. Derived proteins: These are the proteins formed from native protein by
the action of heat, physical forces or chemical factors.

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 A. Simple Proteins
• Major subclasses of simple proteins are as follows:
a) Protamines
• These are small molecules and are soluble in water, dilute acids and
alkalies and non-coagulable by heat.
• They exist as basic proteins in the body.
• They combine with nucleic acids to form nucleoproteins.
• Examples: Salmine, sardinine and cyprinine of fish (sperms) and
testes.

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 A. Simple Proteins

b) Histones

• These are basic proteins, rich in arginine and histidine.

• They are soluble in water, dilute acids and salt solutions but insoluble in ammonia.

• They form conjugated proteins with nucleic acids (DNA).

• Examples: Nucleohistones, chromosomal nucleoproteins and globin of

haemoglobin.

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 A. Simple Proteins

c) Albumins

• These are soluble in water and in dilute salt solutions.

• They are coagulable by heat.

• Examples: Plant albumins: Legumelin in legumes, leucosin in cereals.

• Animal source: Ovalbumin in egg, lactalbumin in milk.

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 A. Simple Proteins
d) Globulins

• Globulins are insoluble in water but soluble in dilute neutral salt solutions.

• They are also heat coagulable.

• Globulins bind with heme, e.g. hemopexin, with metals, e.g. transferrin,
ceruloplasmin and with carbohydrates, e.g. immunoglobulins.

• Examples: In addition to above, ovoglobulin in eggs, lactoglobulin in milk, legumin

from legumes.

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 A. Simple Proteins

e) Gliadins (Prolamines)

• Alcohol soluble plant proteins, insoluble in water or salt solutions.

• Examples: Gliadin of wheat and hordein of barley.

f) Glutelins

• These are plant proteins, insoluble in water but soluble in dilute acids or alkalies.

• They are rich in glutamic acid.

• Examples: Oryzenin of rice and glutelin of wheat.

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 A. Simple Proteins

g) Scleroproteins or Albuminoids

• These are fibrous proteins with great stability and very low solubility
and form supporting structures of animals.

• This group includes keratins, collagens and elastins.

• (a) Keratins: These are characteristic constituents of chidermal tissue

such as horn, hair, nails, wool, hoofs and feathers.
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 A. Simple Proteins

• Human hair has a higher content of cysteine than that of other

species and is called α-keratin.

• β-keratins are deficient in cysteine and, rich in glycine and alanine.

They are present in spider’s web, silk and reptilian scales.

• (b) Collagen: A protein found in connective tissue and bone as long,

thin, partially crystalline substance.
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 A. Simple Proteins

• (c) Elastins: These are the proteins present in yellow elastic fibre of
the connective tissue, ligaments and tendons.

• They are rich in non-polar amino acids such as alanine, leucine, valine
and proline.

• They are formed in large amount in the uterus during pregnancy.

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 B. Conjugated Proteins

• Conjugated proteins are simple proteins combined with a non-protein group

called prosthetic group.

• Protein part is called apoprotein, and entire molecule is called holoprotein.

a) Nucleoproteins

• The nucleoproteins are compounds made up of simple basic proteins such

histone with Nucleic Acids as the prosthetic group.

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 B. Conjugated Proteins

• They are most abundant in tissues having large proportion of nuclear

material such as yeast, thymus and other glandular organs.

• Deoxyribonucleoproteins: It contain DNA as prosthetic group, are

found in nuclei, mitochondria and chloro-plasts.

• Ribonucleoproteins: It occur in nucleoli and ribosome granules. They

have RNA as prosthetic group.
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 B. Conjugated Proteins

b) Mucoproteins or Mucoids

• Mucoproteins are the simple proteins combined with mucopolysaccharides

(MPS) such as hyaluronic acid and the chondroitin sulphate.

• They have hexosamine and hexose sugars as the prosthetic groups.

• Mucoproteins are present in large amounts in umbilical cord.

• Several gonadotropic hormones such as FSH, LH and HCG are mucoproteins.

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 B. Conjugated Proteins
c) Glycoproteins
• Glycoproteins are the proteins with carbohydrate moiety as the
prosthetic group.
• These proteins carry a small amount of carbohydrates( <4%) and
carbohydrate is bound much more firmly in the glycoproteins than the
mucoprotein.
• They carry mannose, galactose, fucose, xylose, arabinose in their
oligosaccharide chains.
• Glycoproteins include mucins, immunoglobulins and many enzymes.
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 B. Conjugated Proteins

d) Chromoproteins

• These are proteins that contain coloured substance as the prosthetic group.

• (i) Haemoproteins: Haem which is a red coloured pigment found as the

prosthetic group in these proteins.

• (ii) Haemoglobin: Respiratory protein found in RB Cells

• (iii) Cytochromes: These are the mitochondrial enzymes of the respiratory chain.

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 B. Conjugated Proteins

e) Catalase: This is the enzyme that decomposes H2O2 to water and O2.

f) Visual purple: is a protein of the retina in which the prosthetic group

is a carotenoid pigment which is purple in colour.

g) Phosphoproteins: These are the proteins conjugated with

phosphoric acid. Casein and Ovovitellin found in milk and eggyolk
respectively are examples.

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 B. Conjugated Proteins

h) Lipoproteins: The lipoproteins are formed in combination with lipids.

HDL, LDL, VLDL and Lecithins are examples.

i) Metalloproteins: As the name indicates, they contain a metal ion as

their prosthetic group. Examples: Ferritin: Contains Fe, Carbonic
Anhydrase: Contains Zn, Ceruloplasmin: Contains Cu.

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 C. Derived Proteins

• This class of proteins are derived from simple or compound proteins by

denaturation or hydrolysis. They are divided in two major groups.

• (a) Primary derived proteins: Denatured or coagulated proteins.

• Heat, X-ray, UV rays, vigorous shaking, acid, alkali can cause denaturation.

• Primary derived proteins are synonymous with denatured proteins in which

peptide bonds remain intact.
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 C. Derived Proteins

• (b) Secondary derived proteins: These are the proteins formed by the
progressive hydrolysis of proteins at their peptide linkages.

• Examples: Protein products obtained by the enzymatic digestion of

proteins. Proteoses, Peptones and Peptides

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 Peptide Linkage and Peptides
• Peptides Are Chains of Amino Acids.

• Polypeptides range in size from small to very large, consisting of two or three to thousands
of linked amino acid residues.

• The –COOH group of one amino is joined to the –NH2 group of another by a covalent bond
called as peptide bond.

• In a peptide, polypeptide, or protein, the amino-terminal end is placed on the left, the
carboxyl-terminal end on the right and the sequence is read left to right, beginning with
the amino-terminal end.

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 Peptide Linkage

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 Peptide Linkage and Peptides

• Three amino acids can be joined by two peptide bonds to form a

tripeptide; similarly, four amino acids can be linked to form a
tetrapeptide, five to form a pentapeptide, and so forth.

• When a few amino acids are joined in this fashion, the structure is
called an oligopeptide. When many amino acids are joined, the
product is called a polypeptide.

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 Peptide Linkage and Peptides

• Proteins may have thousands of amino acid residues.

• The terms “protein” and “polypeptide” are sometimes used

interchangeably but molecules having molecular weights below
10,000 are referred to as polypeptides and those having higher
molecular weights called proteins.

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 Structural organisation of proteins

• Protein structure is normally described at four levels of organisation.

• A. Primary Structure: Primary structure is the linear sequence of amino acids held
together by peptide bonds.

• B. Secondary structure: Refers particularly to stable arrangements of amino acid

residues giving rise to recurring structural patterns (α-helix or β-pleated sheet
structure).

• The bonds holding these patterns are Hydrogen bonds and disulphide bonds.

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 Structural organisation of proteins

• C. Tertiary structure: The polypeptide chain with secondary structure

may be further folded, superfolded and twisted about itself forming
many sizes.

• Such a structural conformation is called tertiary structure.

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 Structural organisation of proteins

• D. Quaternary Structure: Many proteins are made up of only one

peptide chain. However, when a protein consists of two or more
peptide chains held together by non-covalent interactions or by
covalent cross-links, it is referred to as the quaternary structure.

• The assembly is often called as oligomer and each constituent

peptide chain is called as monomer or subunit.

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 Structural organization of proteins

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Denaturation of protiens
• Denaturation is a process in which proteins lose the quaternary structure,
tertiary structure, and secondary structure which is present in their native
state
• by application of some external stress or compound such as
• a strong acid or base,
• a concentrated inorganic salt,
• an organic solvent (e.g., alcohol or chloroform),
• radiation or heat.
• If proteins in a living cell are denatured, this results in disruption of cell
activity and possibly cell death.
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CONTINUED…
• Denatured proteins lose their 3D structure and therefore cannot
function.
• Protein folding is key to whether a globular or membrane protein can
do its job correctly;
• it must be folded into the right shape to function.
• Hydrogen bonds, play a big part in folding.

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COMMON EXAMPLES
• When food is cooked, some of its proteins become denatured.
• This is why boiled eggs become hard and cooked meat becomes firm.
• Cooking the thermally unstable whites turns them opaque, forming an
interconnected solid mass.
• The same transformation can be effected with a denaturing chemical.

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CONTINUED…
• Pouring egg whites into a beaker of acetone will also turn egg whites translucent
and solid.
• Curdled milk is another common example of denatured protein.
• The cold appetizer known as ceviche is prepared by chemically "cooking" raw fish
and shellfish in an acidic citrus marinade, without heat.

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 Protein Denaturants
•Acids
•Acidic protein denaturants include:

•Acetic acid[29]
•Trichloroacetic acid 12% in water
•Sulfosalicylic acid
•Picric acid

•Bases
•Sodium bicarbonate
•Solvents
•Most organic solvents are denaturing, including:

•Ethanol
•alcohol

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Protein Denaturants
• Disulfide bond reducers
• Agents that break disulfide bonds by reduction include:

• 2-Mercaptoethanol
• Dithiothreitol
• TCEP (tris(2-carboxyethyl)phosphine)
• Other
• Mechanical agitation
• Radiation
• Temperature[30]
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Reversible and irreversible denaturation

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EXAMPLE

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