Amino acids & Proteins – an introduction

Proteins are the most abundant biological macromolecules, occurring in all cells and all parts of
cells. Proteins also occur in great variety; thousands of different kinds, ranging in size from
relatively small peptides to huge polymers with molecular weights in the millions, may be found
in a single cell. Moreover, proteins exhibit enormous diversity of biological function and are the
most important final products of the information pathways. Proteins are the molecular
instruments through which genetic informationis expressed.
Relatively simple monomeric subunits provide the key to the structure of the thousands of
different proteins. All proteins, whether from the most ancient lines of bacteria or from the most
complex forms of life, are constructed from the same ubiquitous set of 20 amino acids,
covalently linked in characteristic linear sequences. Because each of these amino acids has a side
chain with distinctive chemical properties, this group of 20 precursor molecules may be regarded
as the alphabet in which the language of protein structure is written. What is most remarkable is
that cells can produce proteins with strikingly different properties and activities by joining the
same 20 amino acids in many different combinations and sequences. From these building
blocks different organisms can make such widely diverse products as enzymes, hormones,
antibodies, transporters, muscle fibers, the lens protein of the eye, feathers, spider webs,
rhinoceros horn, milk proteins, antibiotics, mushroom poisons, and myriad other substances
having distinct biological activities . Among these protein products, the enzymes are the most
varied and specialized. Virtually all cellular reactions are catalyzed by enzymes. Proteins are
polymers of amino acids, with each amino acid residue joined to its neighbor by a specific type
of covalent bond. (The term “residue” reflects the loss of the elements of water when one amino
acid is joined to another.) Proteins can be broken down (hydrolyzed) to their constituent amino
acids by a variety of methods, and the earliest studies of proteins naturally focused on the free
amino acids derived from them. Twenty different amino acids are commonly found in proteins.
The first to be discovered was asparagine, in 1806. The last of the 20 to be found, threonine, was
not identified until 1938. All the amino acids have trivial or common names, in some cases
derived from the source from which they were first isolated. Asparagine was first found in

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 asparagus, and glutamate in wheat gluten; tyrosine was first isolated from cheese (its name is
derived from the Greek tyros, “cheese”); and glycine (Greek glykos, “sweet”) was so named
because of its sweet taste.

Common features of amino acids

All 20 of the common amino acids are α-amino acids. They have a carboxyl group and an amino
group bonded to the same carbon atom (the α carbon) . They differ from each other in their side
chains, or R groups, which vary in structure, size, and electric charge, and which influence the
solubility of the amino acids in water. In addition to these 20 amino acids there are many less
common ones. Some are residues modified after a protein has been synthesized; others are amino
acids present in living organisms but not as constituents of proteins.

The common amino acids of proteins have been assigned three-letter abbreviations and one-letter
symbol, which are used as shorthand to indicate the composition and sequence of amino acids
polymerized in proteins. Two conventions are used to identify the carbons in an amino acid—a
practice that can be confusing. The additional carbons in an R group are commonly designated
β,ᵞ, ᵟ, and so forth, proceeding out from the α carbon. For most other organic molecules, carbon
atoms are simply numbered from one end, giving highest priority (C-1) to the carbon with the
substituent containing the atom of highest atomic number. Within this latter convention, the
carboxyl carbon of an amino acid would be C-1 and the _ carbon would be C-2. In some cases,
such as amino acids with heterocyclic R groups, the Greek lettering system is ambiguous and the
numbering convention is therefore used.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 Stereoisomers
For all the common amino acids except glycine, the α carbon is bonded to four different groups:
a carboxyl group, an amino group, an R group, and a hydrogen atom (Fig. 3–2; in glycine, the R
group is another hydrogen atom). The α carbon atom is thus a chiral center (p. 17). Because of
the tetrahedral arrangement of the bonding orbitals around the α carbon atom, the four different
groups can occupy two unique spatial arrangements, and thus amino acids have two possible
stereoisomers. Since they are nonsuperimposable mirror images of each other (Fig. 3–3), the two
forms represent a class of stereoisomers called enantiomers (see Fig. 1–19). All molecules with
a chiral center are also optically active—that is, they rotate plane-polarized light,
The absolute configurations of simple sugars and amino acids are specified by the D, L system
(Fig. 3–4), based on the absolute configuration of the three-carbon sugar glyceraldehyde, a
convention proposed by Emil Fischer in 1891. (Fischer knew what groups surrounded the
asymmetric carbon of glyceraldehyde but had to guess at their absolute configuration; his guess
was later confirmed by x-ray diffraction analysis.) For all chiral compounds, stereoisomers
having a configuration related to that of L-glyceraldehyde are designated L, and stereoisomers
related to D-glyceraldehyde are designated D. L-Amino acids are those with the α amino group
on the left, and D-amino acids have the α amino group on the right. Historically, the similar l and
d designations were used for levorotatory (rotating light to the left) and dextrorotatory (rotating
light to the right). However, not all L-amino acids are levorotatory, and not all all D-aminoacids
are dextro rotatory.. By Fischer’s convention, L and D refer only to the absolute configuration of
the four substituents around the chiral carbon, not to optical properties of the molecule.
Nearly all biological compounds with a chiral center occur naturally in only one stereoisomeric
form, either D or L. The amino acid residues in protein molecules are exclusively L
stereoisomers. D-Amino acid residues have been found only in a few, generally small peptides,
including some peptides of bacterial cell walls and certain peptide antibiotics.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Classification of amino acids

I. Classification on the basis of R-group

II. Classification on the basis of nutrition

III. Classification on the basis of Catabolism

I) Classification of amino acids on the basis of R-group

Nonpolar, Aliphatic R Groups The R groups in this class of amino acids are nonpolar and
hydrophobic. The side chains of alanine, valine, leucine, and isoleucine tend to cluster together
within proteins, stabilizing protein structure by means of hydrophobic interactions. Glycine has
the simplest structure. Although it is formally nonpolar, its very small side chain makes no real

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 contribution to hydrophobic interactions. Methionine, one of the two sulfur-containing amino
acids, has a nonpolar thioether group in its side chain. Proline has an aliphatic side chain with a
distinctive cyclic structure. The secondary amino (imino) group of proline residues is held in a
rigid conformation that reduces the structural flexibility of polypeptide regions containing
proline.

Classification to be continued in next lecture……………………

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 I) Classification of amino acids on the basis of R-group continued…..

b) Aromatic R Groups :
Phenylalanine, tyrosine, and tryptophan, with their aromatic side chains, are relatively
nonpolar (hydrophobic). All can participate in hydrophobic interactions. The hydroxyl group of
tyrosine can form hydrogen bonds, and it is an important functional group in some enzymes.
Tyrosine and tryptophan are significantly more polar than phenylalanine, because of the tyrosine
hydroxyl group and the nitrogen of the tryptophan indole ring. Tryptophan and tyrosine, and to a
much lesser extent phenylalanine, absorb ultraviolet light This accounts for the characteristic
strong absorbance of light by most proteins at a wavelength of 280 nm, a property exploited by
researchers in the characterization of proteins.

c) Polar, Uncharged R Groups :

The R groups of these amino acids are more soluble in water, or more hydrophilic, than those of
the nonpolar amino acids, because they contain functional groups that form hydrogen bonds with
water. This class of amino acids includes serine, threonine, cysteine, asparagine, and
glutamine. The polarity of serine and threonine is contributed by their hydroxyl groups; that of
cysteine by its sulfhydryl group; and that of asparagine and glutamine by their amide groups.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Asparagine and glutamine are the amides of two other amino acids also found in proteins,
aspartate and glutamate, respectively, to which asparagine and glutamine are easily hydrolyzed
by acid or base. Cysteine is readily oxidized to form a covalently linked dimeric amino acid
called cystine, in which two cysteine molecules or residues are joined by a disulfide bond (Fig.
3–7). The disulfide-linked residues are strongly hydrophobic (nonpolar). Disulfide bonds play a
special role in the structures of many proteins by forming covalent links between parts of a
protein molecule or between
two different polypeptide chains.

Positively Charged (Basic) R Groups The most hydrophilic R groups are those that are either
positively or negatively charged. The amino acids in which the R groups have significant
positive charge at pH 7.0 are lysine, which has a second primary amino group at the _ position
on its aliphatic chain; arginine, which has a positively charged guanidino group; and histidine,
which has an imidazole group. Histidine is the only common amino acid having an ionizable side
chain with a pKa near neutrality. In many enzyme-catalyzed reactions, a His residue facilitates
the reaction by serving as a proton donor/acceptor.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Negatively Charged (Acidic) R Groups:
The two amino acids having R groups with a net negative charge at pH 7.0 are aspartate and
glutamate, each of which has a second carboxyl group.

II. Classification of amino acids on the basis of Nutrition:

1. Essential amino acids:

▪ These amino acids are not synthesized in cells of human beings, so these should be
essentially present in diet.

▪ PVTTIMHALL; Phenylalanine, Valine, Threonine, Tryptophan, Isoleucine, Methionine,

Histidine, Arginine*, Leucine, Lysine

▪ Arginine is conditional amino acids (Essential for infants, non essential for adults)
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 2. Non essential amino acids:

▪ These aminoacids can be synthesized in body, so need not be included in diet.

▪ (GASCAGAGTP); Glycine, Alanine, Serine, Cysteine, Asparagine, Glutamine, Aspartic

acid, Glutamic acid, Tyrosine, Proline

III. Classification of amino acids on the basis of Catabolism

1. Glucogenic amino acids:

▪ These aminoacids serves as precursors of gluconeogenesis for glucose formation
▪ GAMD (Glycine, Alanine, methionine, Aspartic acid).

2. Ketogenic amino acids:

▪ These aminoacids breakdown to form ketone bodies.
▪ Leucine and Lysine

3. Both glucogenic and ketogenic amino acids:

▪ These amino acids breakdown to form precursors for both ketone bodies and glucose.
▪ Isoleucine, Phenylalanine, Tryptophan and tyrosine

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Uncommon Amino Acids/Non standard amino acids
In addition to the 20 common amino acids, proteins may contain residues created by modification of
common residues already incorporated into a polypeptide (Fig. below ). Among these uncommon amino
acids are 4-hydroxyproline, a derivative of proline, and 5-hydroxylysine, derived from lysine. The
former is found in plant cell wall proteins, and both are found in collagen, a fibrous protein of
connective tissues. 6-N Methyllysine is a constituent of myosin, a contractile protein of muscle.
Another important uncommon amino acid is ᵞ carboxyglutamate, found in the bloodclotting protein
prothrombin and in certain other proteins
that bind Ca2_ as part of their biological function. More complex is desmosine, a derivative of four Lys
residues, which is found in the fibrous protein elastin. Selenocysteine is a special case. This rare amino
acid residue is introduced during protein synthesis rather than created through a postsynthetic
modification. It contains selenium rather than the sulfur of cysteine. Actually derived from serine,
selenocysteine is a constituent of just a few known proteins. Some 300 additional amino acids have been
found in cells. They have a variety of functions but are not
constituents of proteins. Ornithine and citrulline deserve special note because they are key
intermediates (metabolites) in the biosynthesis of arginine and in the urea cycle.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Selenocysteine (Sec) and pyrrolysine (Pyl) are rare amino acids that are cotranslationally inserted into
proteins and known as the 21st and 22nd amino acids in the genetic code. Sec and Pyl are encoded by UGA
and UAG codons, respectively, which normally serve as stop signals

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Q. Why do Proteins absorb at 280nm?

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Amino acid Properties

Physical Properties
1. Amino acids are colorless, crystalline solid.
2. All amino acids have a high melting point greater than 200o
3. Solubility: They are soluble in water, slightly soluble in alcohol and dissolve with
difficulty in methanol, ethanol, and propanol. R-group of amino acids and pH of the
solvent play important role in solubility.
4. On heating to high temperatures, they decompose.
5. All amino acids (except glycine) are optically active.

Chemical Properties
1. Amino Acids Can Act as Acids and Bases

A zwitterion is a molecule with functional groups, of which at least one has a positive and one
has a negative electrical charge. The net charge of the entire molecule is zero. Amino acids are
the best-known examples of zwitterions. When an amino acid is dissolved in water, it exists in
solution as the dipolar ion, or zwitterion (German for “hybrid ion”), shown in Figure . A
zwitterion can act as either an acid (proton donor):

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Substances having this dual nature are amphoteric and are often called ampholytes (from
“amphoteric electrolytes”). A simple monoamino monocarboxylic α-amino acid, such as alanine,
is a diprotic acid when fully protonated—it has two groups, the -COOH group and the -NH3 _
group, that can yield protons:

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Chemical Properties (Chemical reactions) of Amino acids Continued…..
The general reactions of amino acids are due to presence of two functional groups namely
carboxyl (-COOH) group and amino (-NH2) group and due to side chains

1. Reactions due to amino group

i) Oxidative deamination-Αn amino group is removed and corresponding α-keto acid is formed.
α -keto acid produced is either converted to glucose or ketone bodies or is completely oxidized.

ii) ii) Transamination-Transfer of an α amino group from an amino acid to an α keto acid to form
a new amino acid and a corresponding keto acid.

iii) Formation of carbamino compound

CO2 binds to α amino acid on the globin chain of hemoglobin to form carbamino hemoglobin

The reaction takes place at alkaline pH and serves as a mechanism for the transfer of Carbon
dioxide from the

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 tissues to the lungs by hemoglobin.

2) Reactions due to carboxyl group

i) Decarboxylation- Amino acids undergo alpha decarboxylation to form corresponding amines.

Examples-

Glutamic acid GABA

Histidine Histamine

Tyrosine Tyramine

ii) Formation of amide linkage

Non α carboxyl group of an acidic amino acid reacts with ammonia by condensation reaction to
form corresponding amides

Aspartic acid→ Asparagine

Glutamic acid → Glutamine

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 3) Reactions due to both amino & carboxyl groups

Formation of peptide bond- Carboxyl group of an amino acid binds with amino group of another
amino acid forming a peptide bond with the loss of one molecule of water.

4. Reactions due to side chains

1) Ester formation
OH containing amino acids e.g. serine, threonine can form esters with phosphoric acid in the
formation of phosphoproteins (figure-1)
OH group containing amino acid can also form: Glycosides – by forming O- glycosidic bond
with carbohydrate residues (figure-2)

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 The Ninhydrin Reaction

In addition to these common reactions of amines and carboxylic acids, common alpha-amino
acids, except proline, undergo a unique reaction with the triketohydrindene hydrate known
as ninhydrin. Among the products of this unusual reaction (shown on the left below) is a purple
colored imino derivative (diketohydrin also known as Ruhemann's purple,) which provides as a
useful color test for these amino acids, most of which are colorless. A common application of the
ninhydrin test is the visualization of amino acids in paper chromatography.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Amino acid + Ninhydrin→ Keto acid + NHr+COz+Hydrindantin
Hydrindantin+ NH3 + Ninhydrin- → Ruhemann's purple

(Note : Proline and hydroxyproline give yellow colour with ninhydrin).

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Introduction to Peptides & Proteins

We now turn to polymers of amino acids, the peptides and proteins. Biologically occurring
polypeptides range in size from small to very large, consisting of two or three to thousands of
linked amino acid residues. Our focus is on the fundamental chemical properties of these
polymers.
Introduction to Peptides and Proteins (Peptides Are Chains of Amino Acids)
Two amino acid molecules can be covalently joined through a substituted amide linkage, termed
a peptide bond, to yield a dipeptide. Such a linkage is formed by removal of the elements of
water (dehydration) from the α-carboxyl group of one amino acid and the α-amino group of
another (Fig). Peptide bond formation is an example of a condensation reaction, a common class
of reactions in living cells. Three amino acids can be joined by two peptide bonds to form a
tripeptide; similarly, amino acids can be linked to form tetrapeptides, pentapeptides, 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.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Proteins may have thousands of amino acid residues. Although the terms “protein” and
“polypeptide” are sometimes used interchangeably, molecules referred to as polypeptides
generally have molecular weights below 10,000, and those called proteins have
higher molecular weights.
As already noted, an amino acid unit in a peptide is often called a residue (the part left over after
losing a hydrogen atom from its amino group and the hydroxyl moiety from its carboxyl group).
In a peptide, the amino acid residue at the end with a free α- amino group is the amino-terminal
(or N-terminal) residue; the residue at the other end, which has a free carboxyl group, is the
carboxyl-terminal (C-terminal) residue. Although hydrolysis of a peptide bond is an exergonic
reaction, it occurs slowly because of its high activation energy. As a result, the peptide bonds in
proteins are quite stable, with an average half-life (t1/2) of about 7 years under most intracellular
conditions.
No generalizations can be made about the molecular weights of biologically active peptides and
proteins in relation to their functions. Naturally occurring peptides
range in length from two to many thousands of amino acid residues. Even the smallest peptides
can have biologically important effects. Consider the commercially synthesized dipeptide L-
aspartyl-L-phenylalanine methyl ester, the artificial sweetener better known as aspartame
or NutraSweet.

Many small peptides exert their effects at very low concentrations. For example, a number of
vertebratehormones are small peptides. These include oxytocin (nine amino acid residues),
which is secreted by the posterior pituitary and stimulates uterine contractions; bradykinin (nine
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 residues), which inhibits inflammation of tissues; and thyrotropin-releasing factor (three
residues), which is formed in the hypothalamus and stimulates the release of another hormone,
thyrotropin, from the anterior pituitary gland. Some extremely toxic mushroom poisons, such as
amanitin, are also small peptides, as are many antibiotics.
Slightly larger are small polypeptides and oligopeptides such as the pancreatic hormone insulin,
which contains two polypeptide chains, one having 30 amino acid residues and the other 21.
Glucagon, another pancreatic hormone, has 29 residues; it opposes the action of insulin.
Corticotropin is a 39-residue hormone of the anterior pituitary gland that stimulates the adrenal
cortex.
Length of the polypeptide chains vary considerably in proteins? Human cytochrome c has 104
amino acid residues linked in a single chain; bovine chymotrypsinogen has 245 residues. At the
extreme is titin, a constituent of vertebrate muscle, which has nearly 27,000 amino acid residues
and a molecular weight of about 3,000,000. The vast majority of naturally occurring proteins are
much smaller than this, containing fewer than 2,000 amino acid residues.
Some proteins consist of a single polypeptide chain, but others, called multisubunit proteins,
have two or more polypeptides associated noncovalently. The individual polypeptide chains in a
multisubunit protein may be identical or different. If at least two are identical the protein is said
to be oligomeric, and the identical units (consisting of one or more polypeptide chains) are
referred to as protomers. Hemoglobin, for example, has four polypeptide subunits: two identical
α- chains and two identical β-chains, all four held together by noncovalent interactions. Each α-
subunit is paired in an identical way with a β subunit within the structure of this multisubunit
protein, so that hemoglobin can be considered either a tetramer of four polypeptide subunits or a
dimer of αβ protomers. A few proteins contain two or more polypeptide chains linked covalently.
For example, the two polypeptide chains of insulin are linked by disulfide bonds. In predominate
such cases, the individual polypeptides are not considconsidered subunits but are commonly
referred to simply as chains.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 We can calculate the approximate number of amino acid residues in a simple protein
containing no other chemical constituents by dividing its molecular weight by 110. Although
the average molecular weight of the 20 common amino acids is about 138, the smaller amino
acids in most proteins. If we take into account the proportions in which the various amino
acids occur in protein the average molecular weight of protein amino acids is nearer to 128.
Because a molecule of water (Mr 18) is removed to create each peptide bond, the average

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Protein Structure

In the previous we discussed several aspect of amino acids, the building block of proteins. Now
in the current lecture, we will discuss more about the protein structure and its function. Proteins
are polymers of amino acids, joined by the covalent bonds, known as pepide bond. A peptide
bond is an amide linkage formed between carboxyl group of first and amino group of second
amino acid with release of water (Figure 32.1, A,B). it is a dehydration synthesis or condensation
reaction. . Every polypeptide chain has a free N- and C terminals For large macromolecules such
as proteins, the tasks of describing and understanding structure are approached at several levels
of complexity, arranged in a kind of conceptual hierarchy. Four levels of protein structure are
commonly defined

✓ Primary Structure
✓ Secondary Structure
✓ Tertiary structure and
✓ Quaternary Structure.

1. Primary structure & Peptide Bond

A description of all covalent bonds (mainly peptide bonds and disulfide bonds) linking amino
acid residues in a polypeptide chain is its primary structure. The most important element of
primary structure is the sequence of amino acid residues. Thus primary structure may be defined
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 as the linear sequence of amino acids in a polypeptide chain that are joined by peptide bonds.
Covalent bonds also place important constraints on the conformation of a polypeptide. In the late
1930s, Linus Pauling and Robert Corey embarked on a series of studies that laid the foundation

for our present understanding of protein structure. They began with a careful analysis of the
peptide bond. The α carbons of adjacent amino acid residues are separated by three covalent
bonds, arranged as Cα-C-N-Cα. X-ray diffraction studies of crystals of amino acids and of
simple dipeptides and tripeptides demonstrated that the peptide CON bond is somewhat shorter
than the CON bond in a simple amine and that the atoms associated with the peptide bond are
coplanar. This indicated a resonance or partial sharing of two pairs of electrons between the
carbonyl oxygen and the amide nitrogen (Fig. 4–2a). The oxygen has a partial negative charge
and the nitrogen a partial positive charge, setting up a small electric dipole. The six atoms of the
peptide group lie in a single plane, with the oxygen atom of the carbonyl group and the
hydrogen atom of the amide nitrogen trans to each other. From these findings Pauling and Corey
concluded that the peptide C-N bonds are unable to rotate freely because of their partial double-
bond character. Rotation is permitted about the N-Cα and the Cα-C bonds. The backbone of a
polypeptide chain can thus be pictured as a series of rigid planes with consecutive planes sharing
a common point of rotation at Cα (Fig. 4–2b). The rigid peptide bonds limit the range of
conformations that can be assumed by a polypeptide chain. By convention, the bond angles
resulting from rotations at Cα are labeled φ (phi) for the N-Cα bond and ψ (psi) for the Cα-C
bond. Again by convention, both φ and ψ are defined as 180º when the polypeptide is in its fully
extended conformation and all peptide groups are in the same plane (Fig. 4–2b). In principle, φ
and ψ can have any value between -180º and +180º, but many values are prohibited by steric
interference between atoms in the polypeptide backbone and amino acid side chains. The
conformation in which both φ and ψ are 0_ (Fig. 4–2c) is prohibited for this reason; this
conformation is used merely as a reference point for describing the angles of rotation. Allowed
values for φ and ψ are graphically revealed when ψ is plotted versus φ in a Ramachandran plot
(Fig. below), introduced by G. N. Ramachandran.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 Protein Structure Continued………

2. Secondary Structure

The secondary structure of a protein is the local fold of the protein backbone. Can a polypeptide
chain fold into a regularly repeating structure? In 1951, Linus Pauling and Robert Corey
proposed two periodic structures called the a helix (alpha helix) and the β pleated sheet (beta
pleated sheet). Subsequently, other structures such as the β turn and omega (Ω ) loop were
identified. Although not periodic, these common turn or loop structures are well defined and
contribute with α helices and βsheets to form the final protein structure. Alpha helices, β strands,
and turns are formed by a regular pattern of hydrogen bonds between the peptide N-H and C=O
groups of amino acids that are near one another in the linear sequence. Such folded segments
are called secondary structure.

i. α helix

In evaluating potential structures, Pauling and Corey considered which conformations of

peptides were sterically allowed and which most fully exploited the hydrogen-bonding capacity
of the backbone NH and CO groups. The first of their proposed structures, the a helix, is a rod
like structure (Figure ). A tightly coiled backbone forms the inner part of the rod and the side
chains extend outward in a helical array. The α helix is stabilized by hydrogen bonds between the
NH and CO groups of the main chain. In particular, the CO group of each amino acid forms a
hydrogen bond with the NH group of the amino acid that is situated four residues ahead in the
sequence (Figure 2.30). Thus, except for amino acids near the ends of an α helix, all the main-
chain CO and NH groups are hydrogen bonded. Each residue is related to the next one by a rise,
also called translation, of 1.5 A along the helix axis and a rotation of 100 degrees, which gives
3.6 amino acid residues per turn of helix. Thus, amino acids spaced three and four apart in the
sequence are spatially quite close to one another in an α helix. In contrast, amino acids spaced
two apart in the sequence are situated on opposite sides of the helix and so are unlikely to make
contact. The pitch of the α helix, which is equal to the product of the translation 1.5 Aº and the
number of residues per turn (3.6), is 5.4 A. The screw sense of a helix can be right-handed
(clockwise) or left -handed (counterclockwise). The Ramachandran diagram reveals that both the
right-handed and the left handed helices are among allowed conformations . However, right -
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 handed helices are energetically more favorable because there is less steric clash between the
side chains and the backbone. Essentially all a helices found in proteins are right-handed. In
schematic representations of proteins, α helices are depicted as twisted ribbons or rods (figure
2.32 ).

Pauling and Corey predicted the structure of the α helix 6 years before it was actually seen in the
x-ray reconstruction of the structure of myoglobin n. The elucidation of the structure of the a
helix is a landmark in biochemistry because it demonstrated that the conformation of a
polypeptide chain could be predicted if the properties of its components are rigorously and
precisely known .

The α helical content of proteins ranges widely, from none to almost 100%. for example, about
75% of the residues in ferritin, a protein that helps store iron, are in IX helices (Figure 2.33).
Indeed, about 25% of all soluble proteins are composed of α helices connected by loops and
turns of the polypeptide chain. Single α helices are usually less than 45 Aº long. Many proteins
that span biological membranes also contain α helices.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 Torsion angle
A measure of the rotation about a bond, usually taken to lie between -180
and +180 degrees. Torsion angles are sometimes called dihedral angles
example ψ &φ.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 Secondary structure contd…………….

Β-Sheets

Pauling and Corey proposed another periodic structural motif, which they named the β- pleated sheet (β because
it was the second structure that they elucidated, the α helix having been the first ). The β pleated sheet (or, more
simply, the β sheet) differs markedly from the rod like α helix. It is composed of two or more polypeptide
chains called β strands. A β strand is almost fully extended rather than being tightly coiled as in the α helix. A
range of extended structures are sterically allowed (Figure 2.34). The distance between adjacent amino acids
along a β strand is approximately 3.5 A, in contrast to a distance of 1. 5 A along an α helix. The side chains of
adjacent amino acids point in opposite directions. A β sheet is formed by linking two or more β strands lying
next to one another through hydrogen bonds. Adjacent chains in a β- sheet can run in opposite directions
(antiparallel β sheet) or in the same direction (parallel β sheet). In the antiparallel arrangement, the NH group
and the CO group of each amino acid are respectively hydrogen bonded to the CO group and the NH group of a
partner on the adjacent chain (Figure 2.36). In the parallel arrangement, the hydrogen-bonding scheme is
slightly more complicated.

For each amino acid, the NH group is hydrogen bonded to the CO group of one amino acid on the adjacent
strand, whereas the CO group is hydrogen bonded to the NH group on the amino acid two residues farther along
the chain (Figure 2.37). The repeat period is shorter for the parallel conformation (6.5 Å, versus 7 Å for
antiparallel) .Many strands, typically 4 or 5 but as many as 10 or more, can come together in β-sheets. Such β-
sheets can be purely antiparallei, purely parallel , or mixed (Figure 2.3H ). In schematic representation β -
strands are usually depicted by broad arrows pointing in the direction of the carboxyl-terminal end to indicate
the type of β- sheet formed parallel or antiparallel.

Some protein structures limit the kinds of amino acids that can occur in the β-sheet. When two or more
Β- sheets are layered close together within a protein, the R groups of the amino acid residues on the touching
surfaces must be relatively small. β-Keratins such as silk fibroin and the fibroin of spider webs have a very high
content of Gly and Ala residues, the two amino acids with the smallest R groups. Indeed, in silk fibroin Gly
and Ala alternate over large parts of the sequence.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 iii) β-turns

In globular proteins, which have a compact folded structure, nearly one-third of the amino acid residues are in
turns or loops where the polypeptide chain reverses direction. These are the connecting elements that link
successive runs of α helix or β conformation. Particularly common are β-turns that connect the ends of two
adjacent segments of an antiparallel β sheet. The structure is a 180º turn involving four amino acid residues,
with the carbonyl oxygen of the residue i forming a hydrogen bond with the amino-group hydrogen of residue
i+3. The peptide groups of the central two residues do not participate in any inte-rresidue hydrogen bonding.
Gly and Pro residues often occur in β-turns, the former because it is small and flexible, the latter because
peptide bonds involving the imino nitrogen of proline readily assume the cis configuration , a form that is
particularly amenable to a tight turn. In other cases, more elaborate structures are responsible for chain
reversals. These structures are called loops or sometimes Ω- loops (omega loops) to suggest their overall shape.
Unlike α- helices and β strands, loops do not have regular, periodic structures. Nonetheless, loop structures are
often rigid and well defin ed . Turns and loops invariably lie on the surfaces of proteins and thus often
participate in interactions between proteins and other molecules. Considerably less common is the γ- turn, a
three residue turn with a hydrogen bond between the first and third.

The term random coil, is used to describe a protein that has lost its secondary structure (the
protein is then said to be denatured).

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
Forces Stablising Protein Structure

Tertiary structure is the three-dimensional structure of a protein. While individual amino

acids in the primary sequence can interact with one another to form secondary structures such as
helices and sheets and individual amino acids from distant parts of the primary sequence can
intermingle via charge-charge, hydrophobic, disulfide, or other interactions, the formation of
these bonds and interactions will serve to change the shape of the overall protein. The folding
that we end up with for a given polypeptide is the tertiary structure.
There are four types of bonding interactions between "side chains" including: hydrogen bonding,
salt bridges, disulfide bonds, and non-polar hydrophobic interactions.

1. Covalent bonds -Disulfide Bridges

Covalent bonds are the strongest chemical bonds contributing to protein structure. Covalent
bonds arise when two atoms share electrons.In addition to the covalent bonds that connect the
atoms of a single amino acid and the covalent peptide bond that links amino acids in a protein
chain, covalent bonds between cysteine side chains can be important determinants of protein
structure. Cysteine is the sole amino acid whose side chain can form covalent bonds, yielding
disulfide bridges with other cysteine side chains: --CH2-S-S-CH2

2. Electrostatic Interactions- Ionic Bonds – (Salt Bridges)

Ionic bonds are formed as amino acids bearing opposite electrical charges are juxtaposed in the
hydrophobic core of proteins. Ionic bonding in the interior is rare because most charged amino
acids lie on the protein surface. Although rare, ionic bonds can be important to protein structure
because they are potent electrostatic attractions that can approach the strength of covalent bonds.

3. Hydrogen Bonds
When two atoms bearing partial negative charges share a partially positively charged hydrogen, the
atoms are engaged in a hydrogen bond (H-bond). The correct 3-D structure of a protein is often
dependent on an intricate network of H-bonds. Individual hydrogen bonds are much weaker
than a covalent bond, but collectively, they can exert strong forces.
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
 These can occur between a variety of atoms, involving:

✓ atoms on two different amino acid sidechains

✓ atoms on amino acid sidechains and water molecules at the protein surface
✓ atoms on amino acid sidechains and protein backbone atoms
✓ backbone atoms and water molecules at the protein surface
✓ backbone atoms on two different amino acids

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 4. Van der Waals Forces

The Van der Waals force is a transient, weak electrical attraction of one atom for another.
Van der Waals attractions exist because every atom has an electron cloud that can fluctuate,
yielding a temporary electric dipole. The transient dipole in one atom can induce a
complementary dipole in another atom, provided the two atoms are quite close. These short-
lived, complementary dipoles provide a weak electrostatic attraction, the Van der Waals
force. Of course, if the two electron clouds of adjacent atoms are too close, repulsive forces
come into play because of the negatively-charged electrons. The appropriate distance
required for Van der Waals attractions differs from atom to atom, based on the size of each
electron cloud, and is referred to as the Van der Waals radius. The dots around atoms in this
and other displays represent Van der Waals radii.Van der Waals attractions, although
transient and weak, can provide an important component of protein structure because of their
sheer number. Most atoms of a protein are packed sufficiently close to others to be involved
in transient Van der Waals attractions.

5. Hydrophobic interactions

The hydrophobic interactions of non-polar side chains are believed to contribute significantly to
the stabilizing of the tertiary structures in proteins. This interaction is really just an application of
the solubility rule that "likes dissolve likes". The non-polar groups mutually repel water and
other polar groups and results in a net attraction of the non-polar groups for each other.
Hydrocarbon alkyl groups on ala, val, leu, and ile interact in this way. In addition, benzene
(aromatic) rings on phe and tyr can "stack" together. In many cases this results in the non-polar

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
 side chains of amino acids being on the inside of a globular protein, while the outside of the
proteins contains mainly polar groups.

Dr. Adfar Yousuf

Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar
Dr. Adfar Yousuf
Assistant professor Dept. of Biotechnology
SP College Srinagar