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Chapter-04 Proteins: Structure and Function

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Chapter 4
Please check highlighted text for correctness.

Proteins:
Structure and Function

Chapter at a Glance
The learner will be able to answer questions on the following topics:

⑧ Peptide bonds Abnormality in protein structure will lead to molecular diseases

with profound alterations in metabolic functions.
⑧ Primary structure of proteins Proteins contain Carbon, Hydrogen, Oxygen and Nitrogen
as the major components while Sulfur and Phos phorus are
⑧ Secondary structure
minor constituents. Nitrogen is characteristic of proteins. On
⑧ Tertiary structure an average, the nitrogen content of ordinary proteins is
16% by weight. All proteins are polymers of amino acids.
⑧ Quaternary structure
Amino Acids are Linked by Peptide Bonds
Alpha carboxyl group of one amino acid reacts with alpha
The word protein is derived from Greek word, “proteios” which amino group of another amino acid to form a peptide bond or
means primary. As the name shows, the proteins are of CO-NH bridge (Fig. 4.1).
paramount importance for biological systems. Out of the total Proteins are synthesized by polymerization of amino acids
dry body weight, 3/4th are made up of proteins. Proteins are through peptide bonds. Two amino acids combined
used for body building; all the major structural and functional
aspects of the body are carried out by protein molecules.
 ⑧ Sequence analysis (study of primary structure) ⑧ unique amino acid sequence decided by the genes. The
primary structure is maintained by the covalent bonds of the
Isoelectric pH of proteins peptide linkages (Fig. 4.1).
⑧ Precipitation reactions of proteins Students should have a clear concept of the term
“sequence”. See the following example:
⑧ Classification of proteins
Gly - Ala - Val (1) Gly - Val - Ala (2)
⑧ Quantitative estimation of proteins Both the tripeptides shown above contain
the same amino acids; but their
sequence is altered. When the sequence
is changed, the peptide is also different.
to form a dipeptide; three amino acids form a tripeptide; four
will make a tetrapeptide; a few amino acids togeth er will
Characteristics of a Peptide Bond
make an oligopeptide; and combination of 10-50 amino acids The peptide bond is a partial double
is a polypeptide. By convention, long poly peptide chains bond. The C–N bond is ‘trans’ in
containing more than 50 amino acids are called proteins. nature and there is no freedom of
In a tripeptide, there are 3 amino acids, but these 3 can rotation because of the partial double
be any of the total 20 amino acids. Thus 20 3 = 8000 different bond character (Fig. 4.2). The distance
permutations and combinations are possible in a tripeptide. Chapter 4: Proteins: Structure and Function 39
An ordinary protein having about 100 amino acids, will have
20100 different possibilities. This number is more than the total
number of atoms present in the whole universe. Thus, even
though there are only 20 amino acids, by changing the
sequence of com
bination of these amino acids, nature produces enor mous
number of markedly different proteins.
Fig. 4.1:
Peptide bond formation

STRUCTURE OF PROTEINS
(ORGANIZATION OF PROTEINS)
Proteins have different levels of structural organization;
primary, secondary, tertiary and quaternary.
Fig.
BOX 4.1: Definitions of levels of organization
4.2: Peptide bond is a partial double bond
1. Primary structure of protein means the order of amino
acids in the polypeptide chain and the location of
disulfide bonds, if any.
2. Secondary structure is the steric relationship of
amino acids, close to each other.
3. Tertiary structure denotes the overall arrangement
and inter-relationship of the various regions, or Fig.
domains of a single polypeptide chain.
4. Quaternary structure results when the proteins 4.3: End groups of polypeptide chain
consist of two or more polypeptide chains held
together by non covalent forces.
Numbering of Amino Acids in Proteins
In a polypeptide chain, at one end there will be one free
alpha amino group. This end is called the amino terminal
Primary Structure (N-terminal) end and the amino acid contri buting the
Sequence of Amino Acids in Proteins alpha-amino group is named as the first amino acid. (Fig.
4.3). Usually the N-terminal amino acid is written on the left
Protein structure is studied as the primary, secondary, hand side when the sequence of the protein is denoted.
tertiary and quaternary levels (Box 4.1). Primary struc ture Incidentally, the biosynthesis
denotes the number and sequence of amino acids in of the protein also starts from the amino terminal end. The
the protein. The higher levels of organization are decided other end of the polypeptide chain is the carboxy
by the primary structure. Each polypeptide chain has a
is 1.32Å which is midway between single GN contributed by the last amino acid (Fig.
bond (1.49Å) and double bond (1.27Å). Ramachandran 1922–2001 4.3). All other alpha amino and alpha
terminal end (C-terminal), where there is carboxyl groups are involved in peptide
The side chains are free to rotate on
a free alpha carboxyl group which is bond formation. Amino
either side of the peptide
bond. The angles of rotation, known as Ramachandran the structural aspects of proteins during 1950s and 1960s).
angles, therefore determine the spatial orientation of the acid residues in polypeptides are named by changing the
peptide chain. (Dr GN Ramachandran did pioneering work on suffix “-ine” to “-yl”, e.g. Glycine to Glycyl. NH2-Gly-Ala-Val-
 COOH In the above example, the amino group of glycine is free;
but carboxyl group of glycine is bonded with amino
40 Textbook of Biochemistry

Fig. 4.4: Primary structure of human insulin

group of alanine; the carboxyl group of Generally the polypeptide chains are cysteine residues of A chain. The species
alanine is, in turn, bonded with the amino linear. However variation is restricted to amino
group of valine; while the carboxyl group together by two interchain disulfide
of valine is free. Therefore this peptide is bonds (Fig. 4.4). The 7th cysteine in A
named as glycyl-alanyl-valine. It is chain and the 7th cysteine in B chain are
connected. Similarly A chain 20th
abbreviated as Gly
cysteine and B chain 19th cysteine are Frederick Sanger NP 1958, 1980
Ala-Val, or. simply as GAV.
connected. There is another intrachain
1918–2013
disulfide bond between 6th and 11th
Branched and Circular Proteins
branching points in the chains may be produced by interchain circulation (Fig. 4.5).
disulphide bridges. The covalent disulphide bonds between
different polypeptide chains in the same protein (interchain) or Primary Structure Determines
portions of the same polypep- Biological Activity
tide chain (intrachain) are also part of the primary structure.
Rarely, instead of the alpha COOH group, the gamma A protein with a specific primary structure will auto matically
carboxyl group of glutamic acid may enter form its natural three dimensional shape. So the higher levels
into peptide bond formation, e.g. Glutathione (gamma of organization are dependent on the primary structure.
glutamyl-cysteinyl-glycine) (see Chapter 18). The term Even a single amino acid change (mutation) in the linear
pseudopeptide is used to denote such a peptide bond sequence may have profound biological effects
formed by carboxyl group, other than that present in alpha
position. Very rarely, protein may be in a circular form, e.g.
Gramicidin.

Primary Structure of Insulin

As an example of the primary structure of a protein, that of
insulin is shown in Figure 4.4. This was originally described by
Sanger in 1955 who received the Nobel Prize in 1958. Insulin
has two polypeptide chains. The A chain (Glycine chain)
has 21 amino acids and B (Phenylalanine) chain has 30
amino acids. They are held
acids in position 8, 9 and 10 in A chain and in C-terminal of B
chain (Fig. 4.4). The amino acid sequence has been
Fig.
conserved to a great extent during evolution.

Proinsulin 4.5: Conversion of proinsulin to active insulin. Arrows = site of action of

proteolytic enzymes
Beta cells of pancreas synthesize insulin as a prohor mone.
Proinsulin is a single polypeptide chain with 86 amino acids. Chapter 4: Proteins: Structure and Function 41
Biologically active insulin (2 chains) is formed by removal of
the central portion of the proinsulin before release. The C-
peptide (connecting peptide) is also released into the
 Linus Pauling Robert B Corey
NP 1954, 1962
1901–1994
Fig. 4.6: Structure of alpha helix 1897–1971
(anti-parallel beta sheet) (Fig. 4.7). Beta
BOX 4.2: Configuration and conformation pleated sheet is the major structural motif in proteins like silk
Fibroin (anti-parallel), Flavodoxin (parallel) and Carbonic
Configuration of a compound denotes the spatial anhydrase (both). Beta bends may be formed in many
relationship between particular atoms, e.g. L- and D- proteins by the abrupt U-turn folding of the chain. Intrachain
amino acids. Conformation means the spatial
relationship of every atom in a molecule, e.g. rotation of
disulfide bridges stabilize these bends.
a portion of the molecule. 42 Textbook of Biochemistry

on the function, e.g. in HbA (normal hemoglobin) the 6th

amino acid in the beta chain is glutamic acid; it is changed to
valine in HbS (sickle cell anemia).

Secondary Structure of Proteins

The term “secondary structure” denotes the configura tional Fig.
relationship between residues, which are about 3–4 amino
acids apart in the linear sequence (Box 4.2). Secondary and
tertiary levels of protein structure are preserved by 4.7: Structure of beta-pleated sheet
noncovalent forces or bonds like hydro gen bonds,
electrostatic bonds, hydrophobic interactions and van der Collagen Helix
Waals forces. These forces are described in Chapter 1.
It is a triple helical structure found in collagen (details in
Alpha Helix Chapter 49).

Pauling (Nobel prize, 1954) and Corey described the alpha Tertiary Structure
helix and beta-pleated sheet structures of polypeptide chains
in 1951. Secondary structure denotes the configurational relation
The alpha helix is the most common and stable ship between residues which are about 3–4 amino acids
conformation for a polypeptide chain. In proteins like apart; or secondary level defines the organization at
hemoglobin and myoglobin, the alpha helix is abundant, immediate vicinity of amino acids. The tertiary structure
whereas it is virtually absent in chymotrypsin. denotes three dimensional structure of the whole protein
(Box 4.1 and Fig. 4.8). The tertiary structure defines the
The alpha helix is a spiral structure (Fig. 4.6). The steric relationship of amino acids which are far apart from
polypeptide bonds form the back-bone and the side chains of each other in the linear sequence, but are close in the
amino acids extend outward. The structure is stabilized by three-dimensional aspect.
hydrogen bonds between NH and C=O groups of the main The tertiary structure is maintained by noncovalent
chain. interactions such as hydrophobic bonds, electrostatic
Each turn is formed by 3.6 residues. The distance bonds and van der Waals forces. The tertiary structure
acquired by native protein is always thermodynamically
between each amino acid residue (translation) is 1.5 Å. The
most stable.
alpha helix is generally right handed. Left handed alpha helix
Examples of different structural motifs are enumer ated
is rare, because amino acids found in proteins are of L-
variety, which exclude left handedness. Proline and in Table 4.1.
hydroxyproline will not allow the formation of alpha helix.
Quaternary Structure
Beta-Pleated Sheet Certain polypeptides will aggregate to form one func
tional protein (Box 4.1 and Fig. 4.8). This is referred to as
The polypeptide chains in beta-pleated sheet is almost fully
the quaternary structure. The protein will lose its function
extended. The distance between adjacent amino acids is
when the subunits are dissociated. The forces that keep
3.5Å. It is stabilized by hydrogen bonds between NH and C=O the quaternary structure are hydrogen bonds, electrostatic
groups of neighboring polypeptide seg bonds, hydrophobic bonds and van der Waals forces.
ments. Adjacent strands in a sheet can run in the same Depending on the number of polypeptide chain, the protein
direction with regard to the amino and carboxy terminal ends may be termed as monomer (1 chain), dimer (2 chains),
of the polypeptide chain (parallel) or in opposite direction tetramer (4 chains) and so on. Each poly peptide chain is
 termed as subunit or monomer.

makes it possible for the precise positioning of the CO2

molecule and the hydroxyl (OH–) ion for the formation of
bicarbonate ion. The enzyme has a zinc ion located at a
deep cleft co-ordinated to histidine residues. The CO2
binding residues are very near to the zinc ion. Water binds
to zinc ion, gets ionized to hydroxyl ion and it binds to the
CO2 which is proximally located. The substrates are brought
in close proximity for the reaction to proceed.

Transport Proteins
Hemoglobin, the transporter of oxygen is a tetrameric
protein (alpha 2, beta 2), with each monomer having a
heme unit (see Chapter 23). Binding of oxygen to one heme
Fig. facilitates oxygen binding by other subunits. Binding of H+
and CO2 promotes release of O2 from hemoglobin. This
allosteric interaction is physiologically important, and is
4.8: Levels of organizations of proteins termed as Bohr effect. Even a single amino acid substitution
alters the structure and thereby the function, e.g. in sickle
cell anemia (HbS), the 6th amino acid in the beta chain is
Homodimer contains two copies of the same polypeptide altered, leading to profound clinical manifestations.
chain. Heterodimer contains two different types of poly
peptides as a functional unit. For example, 2 alpha-chains
Structural Proteins
and 2 beta-chains form the hemoglobin molecule.
Similarly, 2 heavy chains and 2 light chains form one Collagen is the most abundant protein in mammals and is
molecule of immunoglobulin G. Creatine kinase (CK) is a the main fibrous component of skin, bone, tendon, cartilage
dimer. Lactate dehydrogenase (LDH) is a tetramer. and teeth. Collagen forms a superhelical cable where the 3
polypeptide chains are wound around itself (see Chapter
Structure-function Relationship 49). In collagen, every 3rd residue is a glycine. The only
amino acid that can fit into the triple
The functions of proteins are maintained because of their
stranded helix is glycine. Replacement of the central glycine
ability to recognize and interact with a variety of molecules.
by mutations can lead to brittle bone disease. The triple
The three-dimensional structural confor mation provides and
helix of collagen is stabilized by the steric repulsion of the
maintains the functional charac teristics. The three-
rings of hydroxyproline and also by the hydrogen bonds
dimensional structure, in turn, is dependent on the primary
structure. So, any difference in the primary structure may between them. In vitamin C deficiency, failure of
produce a protein which cannot serve its function. To hydroxylation of proline/lysine leads to reduced hydrogen
illustrate the structure function relationship, the following bonding and consequent weakness of collagen (see
three proteins are considered; each belongs to a different Chapter 49). The quarter staggered
class in the functional classification. Chapter 4: Proteins: Structure and Function 43
BOX 4.3: Keratin—Some interesting facts
Enzymes
Mammals contain alpha keratin. It is classified into soft
The first step in enzymatic catalysis is the binding of the and hard keratin depending on the sulfur content. The
enzyme to the substrate. This, in turn, depends on the cysteine residues are responsible for disulfide bridge
structural conformation of the active site of the enzyme, formation which confers characteristic texture for each
which is precisely oriented for substrate binding (see type of the protein. Soft keratin having low sulfur content
is present in skin. Hard keratin is present in hair, horn
Chapter 5). Carbonic anhydrase catalyses the reversible and nails and has high sulfur content. The disulfide
hydration of carbon dioxide. This enzyme bridges resist the forces that try to deform them.
TABLE 4.1: Specific structural motifs in common proteins Springiness of hair is due to the characteristic coiled coil
struc tural motif. When stretched, the coiled coil will
Protein Structural motif untwist and resume the original structure. Hair styling
present like curling and straightening is based on reduction of the
existing disulfide bond and then reoxidation so that new
bonds are formed. Stretching using moist heat breaks
Myoglobin Alpha helix and beta pleated sheet
disulfide bonds. Abnor
malities in keratin structure will cause loss of skin
Collagen Triple helix
integrity and results in diseases like Epidermolysis
bullosa.
Keratin Coiled coil

Elastin No specific motif

Superoxide Antiparallel beta pleated sheet triple helical structure of collagen is responsible for its
dismutase tensile strength. Different arrangements of collagen fibrils
in tissues are seen. Parallel bundles in tendons and sheets
layered at many angles in skin. Heat denatured collagen is determined by mass spectroscopy. Principles of all the
gelatin. See Box 4.3 for details about keratin. above-said techniques are discussed in Chapter 31.

STUDY OF PROTEIN STRUCTURE Steps for Determining the

The first protein to be sequenced was insulin by Sanger in
Primary Structure
1955 (Nobel Prize in 1958). Before studying the struc ture, 1. Determination of the number of polypeptide chains in a
first a pure sample of the protein has to be available. The protein. This is ascertained by treating them with
proteins are extracted and purified by various chro Dansyl chloride, which combines with the N-terminal
matography techniques (ion exchange, adsorption, par amino acid (Fig. 4.9). The tagged polypeptide chains
tition, size exclusion, affinity, HPLC). The purity of the are subjected to complete hydrolysis by boiling with 6
protein thus isolated is studied by electrophoresis (agar, N HCl at 110°C for 18–36 hours under anaerobic
PAGE, iso electric focusing). Further, molecular weight is conditions to give a mixture of amino acids. The
number and nature of
44 Textbook of Biochemistry Carboxypeptidase A and B. These Edman
enzymes specifically hydrolyze and 1916–1977
release the C-terminal amino acid from the
the dansyl amino acids can be determined polypeptide chain. Continued action of the
and will indicate the number of enzyme would release amino acids
polypeptide chains in the protein, e.g. if sequentially from the C-terminal end.
there are two different polypeptide chains Carboxypeptidase A will not act if the C-
in a protein, two different dansyl amino terminal residue is Arginine, Proline or
acids can be identified. Lysine. Carboxypeptidase B will
2. Determination of the amino acid
composition by com plete hydrolysis of the
polypeptide chains, followed by Fig. 4.10: 1-Fluoro-
chromatographic separation and
identification. 2, 4-dinitro-benzene (FDNB)
3. Identification of N-terminal and C-
terminal amino acids (see below).
4. Site specific hydrolysis of the
polypeptide chain using specific enzymes
to get a mixture of over lapping peptides. Fig.4.9: Dansyl
5. Separation and purification of each of
these pep tides, and then analyzing the chloride
amino acid sequence of each of the small Merrifield
peptides, and then deciphering the NP 1984
sequence of the whole protein. 1921–2006

End Group Analysis

The N-terminal amino acid has already
been identified by treatment with dansyl
chloride (Fig. 4.9). Originally Sanger used
Pehr Victor
fluorodinitrobenzene (FDNB, Sanger's
reagent) for identification of N-terminal
amino acid (Fig. 4.10). The C-terminal
amino acid may be identified by
act only if the penultimate residue is Proline. For very long chain proteins, the chain is broken into many
small peptides of overlapping sequences. This is
Sequencing Fig. 4.11: Steps in Edman's degradation process. The numbers show
the amino acid sequence. Amino terminal amino acid reacts with
The purified individual polypeptide chains are then sequenced Edman's reagent; then it is hydrolyzed. This cycle repeats.
using Edman’s degradation technique. Edman’s reagent is
phenyl isothiocyanate. It forms a covalent bond to the N-
terminal amino acid of any peptide chain (Fig. 4.11). This can done by subjecting the polypeptide chain to hydrolysis by two
be identified. The Edman’s reagent would then react with the or more different site specific enzymes. Each of these small
second amino acid which now has the alpha amino group. peptides can be purified and subjected to Edman’s
The degradation technique is useful in sequencing first 10–30 degradation and sequenced.
amino acids. Trypsin hydrolyzes peptide bonds formed by alpha
carboxyl group of Lysine and Arginine.
Partial Hydrolysis
 Chymotrypsin preferentially acts on peptide bonds Shape of the proteins also vary. Thus, Insulin is
formed by carboxyl group of amino acids Phe, Tyr, Trp, or globular, Albumin is oval in shape, while Fibrinogen
Leu. molecule is elongated. Bigger and elongated molecules will
Cyanogen bromide (CNBr) attacks C-side of meth ionine increase the viscosity of the solution.
residue and breaks the peptide bond. Isoelectric pH of amino acids has been described in
Chapter 3. Since proteins are made of amino acids, the pI
of all the constituent amino acids will influence the pI of the
protein. Application of pI is shown in Box 4.4.
Each peptide is then analyzed and the whole seq uence
of the polypeptide is determined as if fitting in the parts of a PRECIPITATION REACTIONS
jigsaw puzzle. The position of disulfide bonds can be OF PROTEINS
determined by cleaving the native protein sample to get
Purification of enzymes and other proteins usually start with
fragments with intact S–S bonds. These fragments are then precipitating them from solution. The stability of proteins in
identified. solution will depend mainly on the charge and hydration.
Polar groups of the proteins (-NH2, COOH, OH groups)
Automatic Sequencing tend to attract water molecules around them to produce a
shell of hydration. Any factor, which neutralizes the
Using the Edman’s degradation technique, sequencing can
charge or removes water of hydration will therefore
be completed within a few hours by automatic machines.
cause precipitation of proteins. The follow ing procedures
Nowadays, DNA sequencing is helping the amino acid are used for protein precipitation:
sequencing. In this method, at first, a rough seq uencing of
protein is done by Edman’s method. Based on this Salting Out
knowledge, small length oligonucleotide primers are made.
These are used to amplify the appropriate gene by When a neutral salt, such as ammonium sulfate or sodium
polymerase chain (PCR) reaction (see Chapter 44) and sulfate is added to protein solution, the shell of hydration is
removed and the protein is precipitated. This is called
correct DNA clone is obtained. The sequenc ing of that part
salting out. As a general rule, higher the molecular weight
of DNA is done. Using the knowledge of the genetic code
of a protein, the salt required for precipitation is lesser.
(see Chapter 41), the sequence of the encoded protein is
Thus globulins are precipitated at half saturation of
identified.
ammonium sulfate; but albu min will need full
Chemical Synthesis of Peptides saturation of ammonium sulfate.

Peptides are artificially synthesized to get pure prepa rations Isoelectric Precipitation
for medical or diagnostic purpose, e.g. HIV anti body in the
blood of AIDS patients is detected by ELISA method. For Proteins are least soluble at their isoelectric pH. Some
this, pure antigen from HIV is to be coated in the test tubes. proteins are precipitated immediately when adjusted to their
Preparation of enough quantity of anti gen from the virus is isoelectric pH. The best example is Casein which forms a
tedious and hazardous. The best flocculent precipitate at pH 4.6 and redissolves in highly
acidic or alkaline solutions. When milk is curdled, the
way is to synthesize the antigenic part of the protein. Bruce
casein forms the white curd, because lactic acid produced
Merrifield in 1961 introduced the solid phase peptide
by the fermentation process lowers the pH to the isoelectric
synthesis (Nobel Prize, 1984). Insulin was the first major
point of casein.
protein chemically synthesized (Katsoyanis, 1964). In
principle, the carboxyl group of the last amino acid is fixed to 46 Textbook of Biochemistry
insoluble polystyrene beads; and other amino acids are BOX 4.4: Significance of isoelectric pH (pI)
added sequentially.
1. The amino acid composition will determine the
isoelectric pH (pI) of protein. The alpha amino group
PHYSICAL PROPERTIES and carboxyl group are utilized for peptide bond
formation, and hence are not ionizable. All other
OF PROTEINS ionizable groups present in the protein will influence
the pI of the protein.
Protein solutions exhibit colloidal properties and there fore
2. At the isoelectric point, the number of anions and
scatter light and exert osmotic pressure. Osmotic pressure cations present on the protein molecule will be equal
of plasma proteins is clinically important (see Chapter 26). and the net charge is zero.
3. At the pI value, the proteins will not migrate in an
Chapter 4: Proteins: Structure and Function 45 electrical field. At the pI, solubility, buffering capacity
and viscosity will be minimum; and precipitation will
be maximum.
Molecular weights of some of the proteins are: Insulin 4. On the acidic side of pI, the proteins are cations and
on alkaline side, they are anions in nature.
(5,700); Hemoglobin (68,000); Albumin (69,000);
5. The pI of pepsin is 1.1; casein 4.6; human albumin 4.7;
Immunoglobulins (1,50,000); Rabbit Papilloma Virus human insulin 5.4; human globulins 6.4; human
Protein (4,70,00,000). hemoglobin 6.7; myoglobin 7; and lysozyme 11.
6. Acidic dyes such as eosin will dissociate into H + + dye
 which will then attach with protein–NH 3+ (protein picrate, etc., and thick flocculent precipitate is formed. In
cations). Basic dyes such as hematoxylin and clinical laboratory phosphotungstic or trichloroacetic acid
methylene blue are dissociated to OH– + dye+, which are usually used for precipitating proteins. Tanning in
will then stain Protein COO– (anions). Thus, the
staining characteristic of a protein is determined by leather processing is based on the protein precipitating
the pI of that protein. effect of tannic acid. Under certain conditions, proteins
undergo denaturation, which is a mild form of precipitation
reaction (Box 4.5). Heat coagulation is an irreversible
precipitation process (Box 4.6 and Fig. 4.13).
Precipitation by Organic Solvents
When an organic solvent is added to the protein solution, CLASSIFICATION OF PROTEINS
water molecules available for proteins are reduced, and
It is almost impossible to correctly classify all proteins. The
precipitation occurs. Organic solvents reduce the dielectric
following classifications are given only to introduce a
constant of the medium which also favors protein
broader idea to the students.
precipitation. Hence alcohol is a powerful protein
precipitating agent. This may explain the disinfectant effect
of alcohol. Classification Based on Functions
1. Catalytic proteins, e.g. enzymes
Precipitation by Heavy Metal Ions 2. Structural proteins, e.g. collagen, elastin 3.
In alkaline medium, proteins have net negative charge, or Contractile proteins, e.g. myosin, actin
are anions. To such a solution, if salts of heavy metals are 4. Transport proteins, e.g. hemoglobin, myoglobin, albu
added, positively charged metal ions can complex with min, transferrin
protein molecules and metal proteinates are precipitated.
Salts of Copper, Zinc, Lead, Cadmium and Mercury are
toxic, because they tend to precipi
tate normal proteins of the gastrointestinal wall. Based on
this principle, raw egg is sometimes used as an antidote
for mercury poisoning.

Precipitation by Alkaloidal Reagents

Tungstic acid, phosphotungstic acid, trichloro acetic
acid, picric acid, sulfosalicylic acid and tannic acid are
powerful protein precipitating agents. These acids Fig.
BOX 4.5: Denaturation of proteins

1. Mild heating, treating with urea, salicylate, X-ray, 4.12: Denaturation of protein
ultraviolet rays, high pressure, vigorous shaking and
similar physico chemical agents produce denaturation.
2. There will be nonspecific alterations in secondary, 5. Regulatory proteins or hormones, e.g. ACTH, insu lin,
tertiary and quaternary structures of protein growth hormone
molecules. Primary structure is not altered during
6. Genetic proteins, e.g. histones
denaturation (Fig. 4.12).
3. In general, during the process the solubility is 7. Protective proteins, e.g. immunoglobulins, inter ferons,
decreased while precipitability of the protein is clotting factors.
increased. It often causes loss of biological activity.
4. Native proteins are often resistant to proteolytic
enzymes, but denatured proteins will have more
Classification Based on Composition
exposed sites for enzyme action. Since cooking leads and Solubility
to denaturation of proteins, cooked foods are more
easily digested. Simple Proteins
5. Denatured proteins are sometimes re-natured when
the physical agent is removed. Ribonuclease is a According to definition, they contain only amino acids. i.
good example for such reversible denaturation. Albumins: They are soluble in water and coagu lated by
Immunoglobulin chains are dissociated when treated heat. Human serum albumin has a mole cular weight of
with urea. When the urea is removed by dialysis, the
subunits are reassociated and biological activity of
69,000, e.g. lactalbumin of milk and egg albumin.
immunoglobulin is regained. ii. Globulins: These are insoluble in pure water, but
6. But many proteins undergo irreversible denaturation. soluble in dilute salt solutions. They are also
For example, albumin cannot be renatured by coagulated by heat, e.g. egg globulin, serum globulins,
removing the physical agent.
legumin of peas.
iii. Protamines: These are soluble in water, dilute acids.
They are not coagulated by heating. They contain large
lower the pH of medium, when proteins carry net positive number of arginine and lysine resi dues, and so are
charges. These protein cations are complexed with strongly basic. Hence they can combine with other
negatively charged ions to form protein-tungstate, protein- acidic proteins. Protamine zinc insulinate is a common
commercial preparation of insulin.
 iv. Prolamins: They are soluble in 70–80% alcohol, but Hemoglobin Globin Heme
insoluble in pure water. They are rich in proline but lack
in lysine, e.g. zein from corn, gliadin of wheat, hordein Nucleoprotein Histones DNA
of barley.
Rhodopsin Opsin 11-cis-retinal
v. Lectins: Lectins are precipitated by 30–60%
ammonium sulfate. They are proteins having high Succinate Protein Riboflavin as
Chapter 4: Proteins: Structure and Function 47 dehydrogenase FAD

BOX 4.6: Significance of heat coagulation Ferritin Apoferritin Iron

When heated at isoelectric point, some proteins will Ceruloplasmin Apoceruloplasmi Copper
denature irreversibly to produce thick floating n
conglomerates called coagulum. This process is called
heat coagulation. Albumin is easily coagulated, and
globulins to a lesser extent. (Fig. 4.13). This is the basis
of ‘Heat and Acetic Acid test’, very commonly employed iii. Nucleoproteins: These are proteins attached to
to detect the presence of albumin in urine (see Chapter
nucleic acids, e.g. Histones. The DNA carries
25).
negative charges, which combines with positively
charged proteins.
iv. Chromoproteins: These are proteins with colored
prosthetic groups. Hemoglobin (Heme, red); Flavo
proteins (Riboflavin, yellow), Visual purple (Vitamin A,
purple) are some examples of chromoproteins.
v. Phosphoproteins: These contain phosphorus.
Casein of milk and vitellin of egg yolk are exam ples.
The phosphoric acid is esterified to the hydroxyl groups
Fig.
of serine and threonine residues of proteins.
vi. Metalloproteins: They contain metal ions. Exam ples
4.13: Heat coagulation. On heating, liquid white portion of egg
are Hemoglobin (Iron), Cytochrome (Iron), Tyro sinase
becomes solid white coagulum
(Copper) and Carbonic anhydrase (Zinc).

affinity to sugar groups. Phytohemagglutinin (PHA), a Derived Proteins

lectin from Phaseolus vulgaris (red kidney bean)
They are degradation products of native proteins. Pro
agglutinates all RBCs and WBCs.
gressive hydrolysis of protein results in smaller and
vi. Scleroproteins: They are insoluble in water, salt
solutions and organic solvents and soluble only in hot smaller chains: Protein → peptones → peptides → amino
strong acids. They form supporting tissues, e.g. acids.
collagen of bone, cartilage and tendon; keratin of hair,
horn, nail and hoof. Classification Based on Shape

Conjugated Proteins Globular Proteins

They are spherical or oval in shape. They are easily
They are combinations of protein with a non-protein part,
soluble, e.g. albumins, globulins and protamines.
called prosthetic group (Table 4.2). Conjugated proteins
may be classified as follows:
i. Glycoproteins: These are proteins combined with
Fibrous Proteins
carbohydrates. Hydroxyl groups of serine or threonine The molecules are elongated or needle shaped. Their
and amide groups of asparagine and glutamine form solubility is minimum. They resist digestion. Collagen,
linkages with carbohydrate residues. Blood group elastin and keratins are examples.
antigens and many serum proteins are glycoproteins.
When the carbohydrate content is more than 10% of Classification Based on Nutritional Value
the molecule, the viscosity is correspondingly Nutritionally Rich Proteins
increased; they are sometimes known as
mucoproteins or proteoglycans. They are also called as complete proteins or first class
ii. Lipoproteins: These are proteins loosely combi ned proteins. They contain all the essential amino
with lipid components. They occur in blood and on cell
membranes. Serum lipoproteins are des cribed in
Chapter 14.
48 Textbook of Biochemistry acids in the required proportion. On supplying these
proteins in the diet, children will grow satisfactorily. A good
TABLE 4.2: Examples of conjugated proteins
example is casein of milk.
Conjugated Protein Prosth
protein part etic Incomplete Proteins
group
 They lack one essential amino acid. They cannot promote Biuret Method
body growth in children; but may be able to sustain the
body weight in adults. Proteins from pulses are deficient Cupric ions chelate with peptide bonds of proteins in alka
in methionine, while proteins of cereals lack in lysine. If line medium produce a pink or violet color. The intensity of
both of them are combined in the diet, adequate growth the color is proportional to the number of peptide bonds.
The color is then compared with a standard pro tein solution
may be obtained. (See mutual supplementation, Chapter
treated with the biuret reagent, and esti mated
35).
colorimetrically. The principle of colorimetry is discussed in
Chapter 31.
Poor Proteins
They lack in many essential amino acids and a diet Advantage
based on these proteins will not even sustain the original
The biuret method is simple one step process, and is the
body weight. Zein from corn lacks tryptophan and lysine.
most widely used method for plasma protein estimations.

Biologically Important Peptides Disadvantage

When 10 or less number of amino acids are joined together,
The sensitivity of the method is less and is unsuitable for
it is called an oligopeptide. Some of them are biologically
estimation of proteins in milligram or microgram quan tities.
active. A few examples are given below:
i. Thyrotropin releasing hormone (TRH) is a tripeptide Lowry’s Method
with the sequence of Glu-His-Pro; but the Glu and Pro
are modified. This is based on the reduction of Folin-Ciocalteau phenol
ii. Glutathione is a tripeptide. It is gamma glutamyl reagent (phosphomolybdic acid and phosphotungstic acid)
cysteinyl glycine (see Chapter 18). It is involved in by the tyrosine and tryptophan residues of protein. A blue
erythrocyte membrane integrity and is important in color is developed, which is compared with that produced by
keeping enzymes in active state. a known standard.
iii. Oxytocin and vasopressin (ADH) are nano peptides;
with 9 amino acids. They are secreted by posterior Advantage
pituitary. Chapter 4: Proteins: Structure and Function 49
iv. Angiotensin I has 10 amino acids and Angiotensin II
has 8 amino acids. They are pressor agents; they
elevate blood pressure. (see Chapter 28). Spectrophotometric Estimation
Polypeptide hormones (more than 10 amino acids) are
described in Chapters 11 and 45. Proteins will absorb ultraviolet light at 280 nm. This is due
to the tyrosine and tryptophan residues in the protein.
Quantitation is done by comparing the absor bance of the
QUANTITATIVE ESTIMATION test solution with a known standard.
Kjeldahl’s Procedure
The protein sample is digested by boiling (360°C) with Advantage
concentrated sulfuric acid in presence of copper sulfate and The method is accurate, simple and highly sensitive up to
sodium sulfate as catalysts. The nitrogen present in the microgram quantities.
protein is reduced to ammonia. The liberated

Nephelometry
It is based on the measurement of scattering of light by
ammonia can be calculated. Then the quantity of nitrogen colloids, originally studied by Richard Zsigmondy (Nobel
originally present in the protein is assessed. Since proteins, Prize, 1925). Nephelometry is defined as the detection of
on an average contain 16% nitrogen, the weight of nitrogen light scattered by turbid particles in solution. If albumin
× 100/16, or nitrogen × 6.25 will give the value of proteins is to be estimated, specific antibody against albumin is
present in the original sample. added. The resultant antigen-antibody com
plex will form turbidity of the solution. A beam of light
Advantage (preferably laser beam) is passed through the solution. The
This is the most accurate and precise method. It is generally particles in the solution will scatter light. The emer gent
used for standardizing a particular protein; that protein is scattered light will be proportional to the turbidity of the
then used for calibrating other proteins employing other solution, which in turn will be proportional to the antigen.
easier methods.
Turbidimetry
Disadvantage
Nephelometry and turbidimetry are based on the same
It takes many days to get the result, and is unsuitable for principle of scattering of light by colloidal particles. In
routine clinical work. nephelometry, emergent light scattered at 60° angle is
observed; while in turbidimetry, light emerging at 180°
angle is detected. Turbidimetry is comparatively cheap
er. In Immunoturbidimetry, the particular protein is pre
cipitated by specific antibody, and the resultant turbidity is
measured. The method is very specific, simple, easy to
perform and cheap.

This method is very sensitive microgram range can be Johan Kjeldahl 1849–1900 Richard A Zsigmondy NP 1925
and protein content in measured. Oliver Howe 1865–1929
Lowry 1910–1996
50 Textbook of Biochemistry peptide bonds.

ELISA Test
7. The primary sequence is genetically determined and is
If proteins of nanogram and picogram quantities are to be unique and fixed for each protein produced by a
estimated, enzyme-linked immunosorbent assay (ELISA) particular species of organism.
technique is to be employed. This is described in detail in 8. Primary structure determines the biological activity of
Chapter 31. the protein. Alterations lead to loss of functional
capacity. E.g. Sickle cell hemoglobin (HbS).
Proteomics 9. The N terminal amino acid is the 1st amino acid having
a free alpha NH2 group and the C terminal amino acid,
Proteomics is the study of the entire galaxy of proteins (the last amino acid) has a free alpha COOH group.
produced by a cell under different conditions. At a 10. A protein having more than one polypeptide chain, like
particular time, a gene is “on” in a particular cell; but it will insulin, has interchain disulfide bonds to hold the
be “off” in another cell. Expression of proteins during chains together. Cysteine forms disulfide link ages
growth and development will be different from the resting between two polypeptide chains in oligomeric proteins.
cell. Proteins produced by a gastro intestinal cell and a 11. Secondary and tertiary levels of structure are
maintained by noncovalent bonds.
neuronal cell will be entirely different. Many proteins
12. The noncovalent bonds maintaining the higher levels of
undergo post-translational modification, that too, at
structure are hydrogen bonds, ionic bonds
different levels at various organs. But genes are the same
(electrostatic bonds), hydrophobic interactions and
in all cells at all times. Therefore study of genes
vander Wal’s forces between the side chains of amino
(genomics) will give only a partial picture of what is going acids.
on in nature. Even though DNA determines the basic 13. The two major types of secondary structure are alpha
genetic structure of an organism, it is the protein which helix and beta pleated sheet structure found in fibrous
actually carries out the body functions. Proteomics aims at proteins. A beta-pleated sheet may further be parallel
studying the protein structure and function. Human body or anti-parallel.
contains hundreds of different cells, which express 14. Tertiary structure of a protein is the most thermo
thousands of proteins, at different times and under the dynamically stable configuration.
influence of different stimuli. Proteomics attempt to study 15. Quaternary structure is present only in certain pro teins
this multifaceted picture in toto. having more than one polypeptide chain eg.
Hemoglobin.
16. The function of a protein is dependent on subunit
LEARNING POINTS CHAPTER 4 interaction.
1. Polymers of amino acids linked by peptide bonds are 17. Chemical properties of the proteins depend on the
called polypeptides. colloidal nature of the particles in solution and the nature of
2. Peptide bonds are covalent bonds between C=O and the side chains. This explains the precipi tation reactions
NH2 groups having a trans configuration. and color reactions of proteins.
18. Solubility of a protein is dependent on the ionic
3. Depending on the number of amino acids, oligopep
concentration of the medium. Hence, proteins may be
tides (10), polypeptides (10-50) and proteins (>50) are
‘salted in’ or ‘salted out’.
formed.
19. Denaturation of protein results in loss of biological
4. Nitrogen content of ordinary proteins is on the average activity but not the primary structure. Denaturation may
16 % by weight. be reversible.
5. Protein structure can be defined and studied at four 20. Proteins can be classified based on (i) Functions (ii)
levels viz. Primary, Secondary, Tertiary and Composition (iii) Shape and (iv) Nutritional value. 21.
Quaternary. Methods of protein estimation include colorimetry (Biuret
6. Proteins have the primary level of structure which and Lowry’s method) and enzyme linked immunosorbent
denotes the linear sequence of amino acids linked by assay (ELISA).
Chapter 4: Proteins: Structure and Function 51

PART-1: ESSAY AND SHORT NOTE QUESTIONS

 4-1. How does the charge on a protein molecule vary with the change in pH of the medium?
4-2. What is end group analysis? What are the reagents used for this purpose?
4-3. Describe the primary, secondary and tertiary structures of proteins. What are the forces, which stabilize them?
4-4. What is the primary structure of a protein? Explain with the help of the structure of Insulin. 4-5. Name two
common types of secondary structures. Mention how they are preserved.
4-6. Name two proteins with quaternary structure. Indicate their subunit make up. How are they maintained? 4-7.
Explain the structural organization of hemoglobin molecule. How does the alteration in amino acid sequence affect
the properties of hemoglobin?
4-8. What are the different techniques used for precipitation of proteins?
4-9. Classify proteins with suitable examples.
4-10. Enumerate the different techniques available for the estimation of proteins. Explain the principle of any one of
them.

SHORT NOTE QUESTIONS

4-11. Primary structure of proteins.
4-12. Alpha helix of proteins.
4-13. Tertiary structure of proteins.
4-14. Quaternary structure of proteins.
4-15. Denaturation of proteins.
4-16. Conjugated proteins.
4-17. Isoelectric pH.
4-18. Isoelectric precipitation.
4-19. Precipitation reactions of proteins.
4-20. Heat coagulation.
4-21. Decarboxylation of amino acids.

PART-2: MULTIPLE CHOICE QUESTIONS

4-1. During denaturation, proteins will not lose their structure, D. The nature of protein folding
with regard to: 52 Textbook of Biochemistry
A. Primary structure
B. Secondary structure
C. Tertiary structure 4-7. One of the following proteins does not have a qua
D. Quaternary structure ternary structure:
4-2. The characteristic features of the peptide bond include all A. Albumin
the following, except: B. Hemoglobin
A. Does not allow freedom of rotation C. Lactate dehydrogenase
B. It is a partial double bond D. Immunoglobulin G
C. Always has cis configuration 4-8. All the following reagents are used for identifying the
D. Absorbs UV light at 280 nm first amino acid in a protein, except:
4-3. The force maintaining the primary structure of a protein: A. Cyanogen bromide
A. Peptide bonds B. Fluorodinitrobenzene
B. Hydrophobic forces C. Dansyl chloride
C. Hydrogen bonds D. Phenyl isothiocyanate
D. Electrostatic (ionic) bonds 4-9. Proteins can be precipitated by the following meth ods,
4-4. The forces maintaining the secondary, tertiary and except:
quaternary structures of a protein are the follow ing, A. Adding alcohol and acetone
except: B. Saturating with ammonium sulphate
A. Electrostatic (ionic) bonds C. Using salts of heavy metals
B. Hydrophobic forces D. Shifting the pH away from the iso electric point
C. Van der Waals forces 4-10. Denatured proteins:
D. Peptide bonds A. Are soluble
4-5. The amino acid which did not allow formation of alpha- B. Are difficult to digest
helix is: C. Are biologically inactive
A. Glutamate D. Peptide bonds are broken
B. Proline 4-11. Which of the following is a simple protein? A.
C. Tyrosine Casein B. Insulin
D. Histidine C. Hemoglobin D. Tyrosinase
4-6. Tertiary structure of a protein describes: A. The 4-12. In glycoproteins, the carbohydrate chains are com
sequence of amino acids bined through glycosidic linkages with:
B. Location of disulphide bonds A. Hydroxyl groups of serine or threonine residues of
C. Amino terminal end amino acid proteins
B. Epsilon amino nitrogen of lysine residues of pro teins
 C. Guanidium group of arginine residues of proteins D. A. Cis double bond
Phenol group of tyrosine residues of proteins B. Isopeptide bond
4-13. The protein which does not answer the aldehyde test
is:
A. Hemoglobin B. Albumin
C. Casein D. Gelatin C. Pseudopeptide bond
4-14. Proteins may be estimated by the following meth ods, D. Peptide bond
except: 4-25. Study of linear sequence of amino acids is done by all
A. Biuret method techniques listed except:
B. Heat coagulation A. End group analysis
C. Kjeldahl’s digestion B. Hydrolysis by proteolytic enzymes
D. Nephelometry C. Analyzing the content of each amino acid
4-15. All the following are examples of tertiary structure of
D. Denaturing the protein
proteins, except:
4-26. Different polypeptide chains are held together by: A.
A. Alpha helix
Peptide bonds B. Disulphide bonds
B. Beta pleated sheet
C. Glycosidic bonds D. Ester bonds
4-27. Primary structure decides:
A. Rate of synthesis of protein

C. Triple stranded helix B. Biological activity of the protein

D. Peptide bonds C. Rate of degradation of the protein
4-16. Tertiary structure of a protein describes: A. D. Effect of proteolytic enzymes on protein
Sequence of amino acids 4-28. Secondary and tertiary levels of protein structure are
B. Location of disulphide bonds dependent on:
C. Amino terminal end amino acid a. Presence of disulfide bonds
D. The nature of protein folding b. Primary structure
4-17. Proteins may be denatured irreversibly by: A. c. PH of the medium
Adding urea d. PK value of component amino acids
B. Bringing to iso electric pH 4-29. The protein having predominantly alpha helical
C. Heat coagulation structure is
D. Reduction with mercaptoethanol A. Collagen
4-18. Lectins are: B. Keratin
A. Animal proteins having specific amino acid binding site
Chapter 4: Proteins: Structure and Function 53
B. Antibody molecules acting against cells
C. Plant proteins having specific carbohydrate bind ing site
D. Blood proteins having a lecithin group C. Fibroin
4-19. Ultraviolet light at 280 nm is absorbed by which
D. Myoglobin
component of proteins?
4-30. Which of the following is NOT true regarding the
A. Peptide bonds
tertiary structure of proteins
B. Sulfhydryl group of cysteine
A. It is a random coil structure
C. Indole ring of tryptophan
B. Disulfide bonds are formed between any two cys teine
D. Imidazole ring of proline
residues
4-20. The nature of the bond linking amino acids to each
other is: C. Position of disulfide bonds are predetermined and fixed
A. Covalent B. Co-ordinate D. Denaturation using reducing agents does not aff ect the
C. Ionic D. Hydrophobic disulfide bonds
4-21. How many peptide bonds are present in glutath ione? 4-31. Protein having a large number of disulfide bonds is: A.
A. 1 B. 2 collagen B. keratin
C. 3 D. 4 C. hemoglobin D. albumin
4-22. Basic difference between two polypeptides is in the — 4-32. Which of the following proteins does not possess a
A. Structural conformation quaternary structure?
B. Primary sequence of amino acids A. Myoglobin
C. Number of side chains B. Lactate dehydrogenase
D. Number of hydrophobic bonds C. Hemoglobin
4-23. Human insulin differs from bovine insulin in: A.
D. Immunoglobulin M
Biological activity
4-33. Which of the following is NOT true regarding hem
B. Number of amino acids
oglobin?
C. Position of disulfide bonds
A. Has 4 independent subunits
D. Sequence of amino acids
B. Each subunit has one heme residue
4-24. A covalent bond between the alpha carboxyl group of
one amino acid and alpha amino group of the C. Each subunit can bind one molecule of oxygen
neighboring amino acid is called D. All four subunits are similar
 ANSWERS OF MULTIPLE CHOICE QUESTIONS
4-1. A 4-2. C 4-3. A 4-4. D 4-5. B 4-6. D 4-7. A 4-8. A 4-9. D 4-10. C 4-11. B 4-12. A 4-13. D 4-14. B 4-15. D 4-16.
D 4-17. C 4-18. D 4-19. C 4-20. A 4-21. C 4-22. B 4-23. D 4-24. D 4-25. D 4-26. B 4-27. B 4-28. B 4-29. A 4-30. D
4-31. B 4-32. A 4-33. D

PART-3: VIVA VOCE QUESTIONS AND ANSWERS

4-1. How proteins are made up of? It is a right handed spiral structure. Each turn is formed by 3.6
Proteins are made by polymerization of amino acids through peptide amino acid residues.
bonds. (Fig. 4.1). 4-18. What is secondary structure of a protein? Secondary
4-2. What is a peptide bond? structure denotes the configurational rela tionship
between residues which are about 3-4 amino acids apart.
Alpha carboxyl group of one amino acid reacts with alpha amino
In other words, secondary level defines the organization
group of another amino acid to form a peptide bond.
at immediate vicinity of amino acids.
4-3. What is a dipeptide?
4-19. What is meant by tertiary structure of a protein? The
Two amino acids are combined to form a dipeptide.
tertiary structure denotes three dimensional struc ture of the
4-4. How many peptide bonds are present in a tripeptide? A
whole protein. It defines the steric relationship
tripeptide is a combination of three amino acids; so there are two
peptide bonds.
4-5. What is a polypeptide?
A combination of 10 to 50 amino acids is called as a polypeptide.
of amino acids which are far apart from each other in the
4-6. What are the levels of organizations of proteins? Proteins
linear sequence.
have primary, secondary, tertiary and quater nary levels of
4-20. What is quaternary structure of a protein? Certain
organization.
polypeptides will aggregate to form one func tional protein. This is
54 Textbook of Biochemistry referred to as the quaternary structure.
4-21. Name proteins having quaternary structure.
Hemoglobin; Immunoglobulins.
4-7. What is meant by primary structure of a protein? It 4-22. What are the reagents that are used for identifying the
denotes the number and sequence of amino acids in the protein. first amino acid in a protein?
4-8. What force maintains the primary structure? The Fluorodinitrobenzene; Dansyl chloride; Phenyl iso thio cyanate.
primary structure is maintained by the covalent bonds of the 4-23. What is isoelectric point of a protein? At the isoelectric
peptide linkages. point, the number of anions and cati ons present on the protein
4-9. How are the end amino acids of proteins called? The molecule will be equal and the netcharge is zero.
end where there is a free alpha amino group, is called the amino
4-24. What are the features of isoelectric point? At the pI
terminal (N-terminal) end. The other end of the polypeptide chain
value, the proteins will not migrate in an elec trical field; solubility,
is called the carboxy ter minal end (C-terminal), where there is a
buffering capacity and viscosity will be minimum and precipitation
free alpha car boxyl group.
will be maximum. 4-25. What is the isoelectric pH of human
4-10. What is a pseudopeptide?
albumin? It is 4.7
The pseudopeptide is a peptide bond formed by car boxyl group,
4-26. How proteins are precipitated from solution? Any factor
other than that of alpha position.
which neutralizes the charge or removes water of hydration will
4-11. Can you give an example of a pseudopeptide?
cause precipitation of proteins. 4-27. How is albumin
Glutathione (gamma-glutamyl-cysteinyl-glycine). 4-12. What are
precipitated?
the salient structural features of insulin? It has two
By Full saturation of ammonium sulphate. 4-28. What
polypeptide chains with 51 amino acids. Chain A has 21 amino
about gobulins?
acids and Chain B has 30 amino acids. The two chains are held
together by disulfide bridges. Globulins are precipitated by half saturation of ammo nium
4-13. What is proinsulin? sulphate.
Insulin is synthesized by the beta cells of pancreas as a single 4-29. Give example of isoelectric precipitation. Casein is
polypeptide chain with 86 amino acids. The middle part is precipitated when the solution is brought to isoelectric pH.
removed as C peptide; the remaining part becomes A 4-30. What is the isoelectric pH of casein?
and B chains. It is 4.6.
4-14. What is a mutation? 4-31. What are the features of denaturation? The secondary,
Amino acid change in the linear sequence is called a mutation. tertiary and quaternary structures are lost; but primary structure is
4-15. Can you give an example? preserved. The functional activity is lost. Denatured proteins are
Sickle cell anemia due to hemoglobin S (HbS). 4-16. What are insoluble and easily precipitated.
the forces that maintain the secondary, tertiary and 4-32. What is heat coagulation?
quaternary structures of a protein? Hydrogen bonds; When heated at isoelectric point, some proteins will denature
Electrostatic bonds; van der Waal's forces and Hydrophobic irreversibly to produce thick floating conglom erates called
bonds. coagulum. This is called heat coagulation.
4-17. What are the salient features of alpha structure of 4-33. Give examples of proteins that coagulate. Albumin is
proteins? easily coagulated, and globulins to a lesser extent.
 4-34. How are proteins classified? Casein.
They may be classified (a) depending on the function 4-42. Why some proteins are nutritionally poor? They lack
(b) based on the physicochemical characteristics or (c) in many essential amino acids and a diet based on these
based on their nutritional value. proteins will not even sustain the body weight.
4-35. How are proteins classified on physical basis? 4-43. Give an example of nutritionally poor protein. Zein
Simple proteins, conjugated proteins and derived pro teins. from corn lacks tryptophan and lysine. 4-44. What is the
advantage of biuret method? The biuret method is simple
4-36. Give examples of simple proteins.
one step process, and is the most widely used method for
Albumins, Globulins, Protamines, Prolamins, Lectins,
plasma protein esti mations.
Scleroproteins.
4-45. What is the disadvantage of biuret method? The
4-37. Give examples of scleroproteins. sensitivity of the method is less and is unsuitable for estimation
Collagen of bone, cartilage and tendon; keratin of hair. 4.38. of proteins in milligram or microgram quantities.
What are conjugated proteins?
Combinations of protein with a non-protein part, called
prosthetic group.
4-39. How are conjugated group subclassified?
Glycoproteins, lipoproteins, nucleoproteins, chromo proteins,
phospho-proteins and metalloproteins.

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Chapter 4: Proteins: Structure and Function 55

4-40. Give some examples of chromoproteins

Hemoglobin; Flavoproteins, Visual purple.
4-41. Give an example of a nutritionally rich protein (first
class protein).