STRUCTURE OF PROTEINS

The structure of proteins is much more complex than that of simple organic molecules. Many protein molecules
consist of a chain of amino acids twisted and folded into a complex three-dimensional structure. The complex 3D
structures of proteins impart unique features to proteins that allow them to function in diverse ways.
There are four levels of organization in proteins structure: primary, secondary, tertiary, and quaternary.

Primary Structure of Protein

The primary structure of a protein is the linear sequence of the side chains that are connected to the protein
backbone.

Fig 1a The primary Structure of Protein

Each protein has a unique sequence of amino acid residues that cause it to fold into a distinctive shape that allows
the protein to function properly.
Peptide bonds
Covalent bonds place important constraints on the conformation of a polypeptide. In the late 1930s, Linus Pauling
and Robert Corey embarked 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 C-N bond is somewhat shorter than the C-N 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 as shown below:

Fig. 1b The partial sharing of electrons in peptide bond

Generally, protein structure is stabilized by multiple weak interactions. Hydrophobic interactions are the major
contributors to stabilizing the globular form of most soluble proteins; hydrogen bonds and ionic interactions are
optimized in the specific structures that are thermodynamically most stable. The nature of the covalent bonds in the
polypeptide backbone places constraints on structure. The peptide bond has a partial double bond character that
keeps the entire six-atom peptide group in a rigid planar configuration.
The N-Cα and Cα-C bonds can rotate to assume bond angles of ɸand ψ, respectively.

Secondary Structure of Protein

The term secondary structure refers to the local conformation of some part of a polypeptide. This is a common
regular folding patterns of the polypeptide backbone. It is the spatial arrangement of amino acid residues that are
adjacent in the primary structure.
Pauling and Corey predicted the existence of these secondary structuresin 1951, several years before the first
complete protein structure was elucidated. Hydrogen bonding causes protein chains to fold and align to produce
orderly patterns called secondary structures. A few types of secondary structure are particularly stable and occur
widely in proteins. The most prominent are the α-helix and β-conformations.

The α-Helix of a Secondary Protein Structure

The simplest arrangement the polypeptide chain could assume with its rigid peptide bonds (but other single bonds
free to rotate)is a helical structure, which Pauling and Corey called the α-helix. The α-helix is held in this shape by
hydrogen bonding interactions between amide groups, with the side chains extending outward from the coil. The
amount of α-helix coiling in proteins is highly variable.

Hydrogen bond between the C=O bond and N-H bond

In the α-helix structure the polypeptide backbone is tightly wound around an imaginary axis drawn longitudinally
through the middle of the helix, and the R groups of the amino acid residues protrude outward from the helical
backbone. More generally, about one-fourth of all amino acid residues in polypeptides are found in α-helices, the
exact fraction varying greatly from one protein to the next. The α-helix form readily than the other possible
conformations because it makes optimal use of internal hydrogen bonds. The structure is stabilized by a hydrogen
bond between the hydrogen atom attached to the electronegative nitrogen atom of a peptide linkage and the
electronegative carbonyl oxygen atom of the fourth amino acid on the amino-terminal side of that peptide bond. All
the hydrogen bonds in the α-helix combined give the entire helical structure considerable stability. Further model-
building experiments have shown that an α-helix can form in polypeptides consisting ofeither L- or D-amino acids.
However, all residues must be of one stereoisomeric series.
Fig. 4 The α-helix structure of protein
Negatively charged amino acids are often found near the amino terminus of the helical segment, where they have a
stabilizing interaction with the positive charge of the helix dipole; a positively charged amino acid at the
aminoterminal end is destabilizing. The opposite is true at the carboxyl-terminal end of the helical segment.
There are five different kinds of constraints affecting the stability of an α-helix namely:
(1) the electrostatic repulsion (or attraction) between successive amino acid residues with charged R groups,
(2) the bulkiness of adjacent R groups,
(3) the interactions between R groups spaced three (or four) residues apart,
(4) the occurrence of Proline and Glycine residues, and
(5) the interaction between amino acid residues at the ends of the helical segment and the electric dipole inherent to
the α-helix.

Fig. 5 The helix dipole

The tendency of a given segment of a polypeptide chain to fold up as an α-helix therefore depends on the identity
and sequence of amino acid residues within the segment. Pauling and Corey predicted a second type of repetitive
structure, the β-conformation.

The β-Conformation
This is a more extended conformation of polypeptide chains, and its structure has been confirmed by x-ray analysis.
In the β-conformation, the backbone of the polypeptide chain is extended into a zigzag rather than helical structure.
The zigzag polypeptide chains can be arranged side by side to form a structure resembling a series of pleats. In this
arrangement, called a β-sheet, hydrogen bonds are formed between adjacent segments of polypeptide chain. The
individual segments that form a β-sheet are usually nearby on the polypeptide chain, but can also be quite distant
from each other in the linear sequence of the polypeptide; they may even be segments in different polypeptide
chains. The R groups of adjacent amino acids protrude from the zigzag structure in opposite directions, creating
the alternating pattern seen in the side views in Fig. 6 below.
Figure 6 The antiparallel view of β-pleated sheet of the protein structure

The adjacent polypeptide chains in a β-sheet can be either parallel or antiparallel (having the same or opposite
amino-to-carboxyl orientations, respectively). The structures are somewhat similar, although the repeat period is
shorter for the parallel conformation (6.5 Å, versus 7 Å for antiparallel) and the hydrogen bonding patterns are
different. Examples include β-Keratins such as silk fibroin and the fibroin of spider webs.
β-Turns
β-turns occur frequently whenever strands in β-sheets change the direction, the 180° turn is accomplished over four
amino acids which is stabilized by a hydrogen bond from a carbonyl oxygen to amide proton three residues down
the sequence. Proline in position 2 or glycine in position 3 is common in β turns

Secondary structure is the regular arrangement of amino acid residues in a segment of a polypeptide chain, in
which each residue is spatially related to its neighbors in the same way. The most common secondary structures are
the α-helix, the β-conformation, and β-turns. The secondary structure of a polypeptide segment can be completely
defined if the ɸ and ψ angles are known for all amino acid residues in that segment.

Tertiary and Quaternary Structures of Proteins

Tertiary Structure of Proteins
The overall three-dimensional arrangement of all atoms in a protein is referred to as the protein’s tertiary structure.
The tertiary structure of a protein refers to the bending and folding of the protein into a specific three-dimensional
shape. The bending and folding of the tertiary structure of a protein into specific three-dimensional shape involves
four types of interactions between the R side chains of the amino acids residues:

1. Disulfide bridges can form between two cysteine residues that are close to each other in the same chain, or
between cysteine residues in different chains. These bridges hold the protein chain in a loop or some other 3D shape.
2. Salt bridges are attractions between ions that result from the interactions of the ionized side chains of acidic
amino acids (—COO-) and the side chains of basic amino acids (—NH3+).
3. Hydrogen bonds can form between varieties of side chains, especially those that contain:
Hydrogen bonding also influences the secondary structure, but here the hydrogen bonding is between R groups,
while in secondary structures it is between the C=O and NH portions of the backbone.
4. Hydrophobic interactions result from the attraction of nonpolar groups, or when they are forced together by
their mutual repulsion of the aqueous solvent. These interactions are particularly important between the benzene
rings in phenylalanine or tryptophan. This type of interaction is relatively weak, but since it acts over large surface
areas, the net effect is a strong interaction.

Figure 7 The forces in the tertiary structure of protein

Tertiary structure includes longer-range aspects of amino acid sequence. Amino acids that are far apart in the
polypeptide sequence and that reside in different types of secondary structure may interact within the completely
folded structure of a protein. The location of bends (including β-turns) in the polypeptide chain and the direction and
angle of these bends are determined by the number and location of specific bend-producing residues, such as
proline, threonine, serine, and glycine. Interacting segments of polypeptide chains are held in their characteristic
tertiary positions by different kinds of weak bonding interactions (and sometimes by covalent bonds such as
disulfide cross-links) between the segments.
The tertiary structure involves the folding of the helices of globular protein, it is the spatial arrangement of amino
acids that are far apart in linear sequence and to the pattern of disulfide bonds. The dividing line between the
secondary and tertiary structure is a matter of taste. Myoglobin is an example of tertiary structure protein with a 3-D
structure, x-ray crystallographic studies have revealed about 300 proteins with 3-D structure.
Myoglobin
This contains a single polypeptide chain of 153 amino acid residues and a single prosthetic iron-porphyrin (or heme)
group, identical with hemoglobin. Myoglobin is abundant in the muscles of dividing mammals such as whale, seal
and porpoise. Storage of oxygen by muscle myoglobin permits these animals to remain submerged under water for
long periods. The function of myoglobin is to bind oxygen in the muscles and enhance its transport to the
mitochondria, which consume it during respiration.
Figure 8: Structure of Myoglobin

Quaternary Structure of Protein

When two or more polypeptide chains are held together by disulfide bridges, salt bridges, hydrogen bond, or
hydrophobic interactions, forming a larger protein complex, a quaternary structure is formed. Each of the
polypeptide subunits has its own primary, secondary, and tertiary structure. The arrangement of the subunits to form
a larger protein is the quaternary structure of the protein. In order words, quaternary structure refers to the spatial
arrangement of subunits and the nature of their contact. In considering these higher levels of structure, it is useful to
classify proteins into two major groups namely:

1. fibrous proteins, having polypeptide chains arranged in long strands or sheets, and
2. globular proteins, having polypeptide chains folded into a spherical or globular shape.
The two groups are structurally distinct: fibrous proteins usually consist largely of a single type of secondary
structure, while globular proteins often contain several types of secondary structure.
The two groups differ functionally in that the structures that provide support, shape, and external protection to
vertebrates are made of fibrous proteins, whereas most enzymes and regulatory proteins are globular proteins.
Certain fibrous proteins played a key role in the development of our modern understanding of protein structure and
provide particularly clear examples of the relationship between structure and function.

An example of a quaternary structure protein is hemoglobin, which is a tetrameric protein that contains 4
polypeptide chains held together by non-covalent interactions. The adult hemoglobin consist of two alpha (α) chains
and two beta (β) chains. Hemoglobin has a molecular weight of 64,500, 4 times as large as myoglobin.
Max Perutz, John C. Kendrew and their colleagues in 1959 determined the 3-D structure of hemoglobin. This
consists of two α and β chains; the α-chain has valine at the N-terminal and arginine at the C-terminal whereas the β-
chain has valine situated at the N-terminal and histidine at the C-terminal. The α- and β-chain are held together as a
pair by ionic and hydrogen bonds, hence the four polypeptide fit together almost tetrahedrally to produce the
characteristic quaternary structure.
Figure 9: Quaternary structure of hemoglobin molecule