Structure organization of the Proteins includes: 1. Primary structure 2. Secondary Structure 3. Tertiary Structure 4. Quaternary Structure

1. Rahul SIR
Lecturer, Department of Med-Surg
Co-chair of the South East Asia Regional Hub within the Challenger’s
Committee, Nursing Now Challenge, London, UK
Structure of Proteins
Bio-Chemistry

2. Specific learning objectives
🠶Structure organization of the Proteins includes:
1) Primary structure
2) Secondary Structure
3) Tertiary Structure
4) Quaternary Structure
🠶 Enumerate and describe pathophysiological conditions arising due
to defective protein structure
🠶 Describe structure of collagen and its structure-function relationship with clinical
correlation
🠶 Describe the structure of Myoglobin and Hemoglobin and its structure-function
relationship

3. Structural organization of Proteins
Fig.3.16. Lehninger Principles of Biochemistry

4. Protein’s conformation stabilized by mainly weak interactions
🠶 In protein structure, stability defined as tendency to maintain a native
conformation
🠶 Protein conformation with lowest free energy (most stable) is the one with
maximum number of weak interactions
🠶 Native proteins are marginally stable; ΔG separating the folded and
unfolded states in proteins under physiological conditions is in range of
20-65kJ/mol

5. Cont--
• For all proteins, weak interactions imp in folding of polypeptide chain into
2° and 3° structure, association of multiple polypeptides to form
4°structure
• Net stability contributed by hydrogen bond, or the difference in free
energies of the folded and unfolded states, may be close to zero
🠶 Protein stability depends on hydrophobic interactions: Hydrophobic aa
side chains tend to cluster in protein’s interior, away from water.
• AA seq of most proteins contains hydrophobic aa side chains, positioned
so that form clusters when protein fold, forming a hydrophobic protein
core.

6. Primary Structure
🠶Amino acids (aa) are linked by peptide bonds to form polypeptide
chains.
🠶Ordered sequence of aa in a polypeptide chains is primary structure
of protein.
🠶It is unique primary structure that enables a polypeptide chain to fold
into a specific 3-D structure that gives protein its chemical and
physiological properties.

7. Amino Acids are Polymerized into Peptides and Proteins
 Condensation reaction: α-carboxyl group of
one aa with side chain R1 forms a covalent
peptide bond with α-amino group of other aa
with side chain R2 by elimination of a
molecule of H2O
 Exergonic reaction: Ex. Hydrolysis of a
peptide bond, it occurs slowly because it has
a high activation energy, so peptide bond
become quit stable, with average half lifḛ
7yrs under intracellular conditions
Fig3.13: Lehninger Principles of Biochemistry by David L Nelson, 6th Ed

8. Cont--
 Dipeptide (contain two aa bonded to each
other via a single peptide bond)
 Tripeptides (contains three aa) form a
second peptide bond through its terminal
carboxylic acid group and amino of a third aa
(R3).
 Repetition of this process form a polypeptide
or protein of specific aa sequence
(R1R,2R,3R4∙∙ ∙Rn). Fig. 2.8. Peptide bond formation:
Textbook of Biochemistry with Clinical Correlations
4th edition by Thomas M Devlin

9. Components of a Polypeptide Chain
 It consists of a repeating backbone part,
and a variable part consists distinctive
side chains.
 Polypeptide backbone is rich in hydrogen-
bonding. Each residue contains a
carbonyl group (exception of proline, an
NH group)
 Mass of a protein expressed in units of
daltons; one Dalton (Da) is equal to one
atomic mass unit.
Fig.2.15: Components of a polypeptide chain: Biochemistry 7th
edition by Berg, Tymoczko and Stryer

10. Peptide Bond has Partial Double-Bond Character
 Single bond linked α-carboxyl and α-nitrogen
atoms, this peptide bond exhibits partial
double-bond character.
 Bond that connects a carbonyl carbon to an α-
nitrogen cannot rotate and thus conformation
of peptide backbone is constrained.
 Almost all peptide bonds in proteins are trans.
 Therefore, O, C, N, and H atoms of a peptide
bond are coplanar.
Fig.2.18: Peptide bonds are planar: Biochemistry 7th edition by Berg, Tymoczko and Stryer

11. Rotation about Bonds in a Polypeptide
 Peptide bond conformation defined by values
of Phi (φ) and psi (ψ) angle or dihedral angles
 φ is angle of rotation about bond between N-Cα
bond
 ψ is angle of rotation about bond between Cα-C
bond
 Both φ and ψ defined as ±180° when
peptide
polypeptide fully extended and all
groups in same plane
Fig.2.22: Biochemistry 7th edition by Berg, Tymoczko and Stryer
α

12. Cont--
 Dihedral angles increases as the distal (4th) atom
is rotated clockwise
 From ±180° position, dihedral angle increases
from -180° to 0°, at which point 1st and 4th atoms
eclipsed
 Rotation continue from 0° to +180° to bring
structure back to starting point
 φ and ψ have any value bet -180° to +180°, but
many values prohibited by steric hindrance bet
atoms in polypeptide backbone and aa side
chains
Fig.4.2 (d). Lehninger Principles of Biochemistry, 6th Ed.

13. Ramachandran Plot
🠶 Ramachandran plot introduced by G.N.Ramachandran in 1963
🠶 Give visual description of combinations of φ and ψ angles that permitted
in peptide backbone or not permitted due to steric hindrances
🠶 Allowed values for φ and ψ become evident when ψ is plotted versus φ
in a Ramachandran plot
🠶This plot are useful tools used to test the quality of 3-D protein
structures deposited in international databases
🠶In case of Gly residue, less sterically hindrance, has much broader
ranges of allowed confirmations

14. Cont--
 φ and ψ values bet. -180° and +180°, but
many values are prohibited by steric
interference between atoms in polypeptide
backbone and aa side chains
 Allowed/favorable regions with no steric
overlap are shown in dark green; borderline
regions are shown in light green
 In peptides, for aa other than glycine, most
combinations of φ and ψ angles are
disallowed due to steric hindrance
Fig.2.23: Biochemistry 7th edition by Berg, Tymoczko and Stryer

15. Amino Acid Sequences have Direction
 A polypeptide chain has polarity because
its ends are different: an α-amino group is
present at one end and an α-carboxyl
group at the other.
 Amino end is taken to be beginning of a
polypeptide chain, and so sequence of aa
in a polypeptide chain is written starting
with amino-terminal residue.
Fig.2.14: Amino acid sequences have direction: Biochemistry 7th
edition by Berg, Tymoczko and Stryer

16. Secondary Structure of Proteins
🠶 Regular arrangements of protein chain are stabilized by hydrogen
bonding
🠶 Polypeptide chains fold into regular structures such as alpha (α) helix,
beta (β) sheet, and turns and loops.
🠶 α-helices, β-strands, and turns are formed by a regular pattern of
hydrogen bonds between peptide N-H and C=O groups of aa that are
near one another in linear sequence. Such folded segments are called
secondary structure.

17. Alpha Helix Is a Coiled Structure Stabilized by Intrachain Hydrogen Bonds
 α-helix is stabilized by intrachain hydrogen
bonding between CO and NH groups along
parallel to helical turn.
 R groups of each aminoacyl residue in an α-
helix face outward.
 Pitch of α-helix: length of one complete turn
along helix axis and is equal to product of rise
(1.5 Å) and number of aa per turn (3.6), or 5.4
Å.
Fig.4.4 b: Ball-and-stick model of a right handed α helix. Lehninger Principles
of Biochemistry
Amino terminus

18. Cont--
 Largely α-helical protein: Ferritin
 ~75% of residues in ferritin, a protein that
helps store iron, are in α-helices.
 ~25% of all soluble proteins are composed
of α-helices connected by loops and turns
of polypeptide chain.
 Many proteins that span biological
membranes also contain α-helices.
Fig.2.28: Biochemistry 7th edition by Berg, Tymoczko and Stryer

19. Type of constraints affect stability of α-helix
🠶Intrinsic susceptibility of aa residue to form α-helix
🠶 Interaction bet R groups (spaced 3 or 4 residues apart)
🠶Bulkiness of adjacent R groups
🠶Occurrence of Pro and Gly residues
🠶 Interaction bet aa residues at end of helical segment and electric dipole
inherent to α-helix

20. β-Sheets Stabilized by Hydrogen Bonding Between Polypeptide Strands
 Backbone of polypeptide chain
extended into zigzag
 Arrangement of many segments side
by side in β-conformation is called β-
sheets
 Hydrogen bonds form bet adjacent
segments of polypeptide chain within
sheet
 Side chains are above and below
plane of strands.
Fig.2.30: Biochemistry 7th edition by Berg, Tymoczko and Stryer
C
O
Cα
N
H

21. Parallel β-sheet
 In parallel arrangement, for each aa,
NH group is hydrogen bonded to CO
group of one aa on adjacent strand
 1:2 H-bond pattern: 1 a.a bonds with 2
other a.a in an opposing stand.
Fig.2.32: Biochemistry 7th edition by Berg, Tymoczko and Stryer
N
H
O
C
Cα
R
O
C

22. Antiparallel β-sheet
Fig.2.31: Biochemistry 7th edition by Berg, Tymoczko and Stryer
N
H
C
O
Cα
 Adjacent chains in a β sheet can run
in opposite directions (antiparallel β
sheet) or in same direction (parallel β
sheet).
 Follow 1:1 H-bond pattern
 In antiparallel arrangement, NH group
and CO group of each aa are
respectively hydrogen bonded to CO
group and NH group of a partner on
adjacent chain.
H
N C
o

23. Reverse Turns
 In globular proteins, has compact folded
structure, some aa residues are in turns or
loops where polypeptide chain reverses
direction. Turns connect helical twists and
sheets.
 Structure is a 180° turn involving four aa
residues, with CO of residue i or 1st residue
forming a hydrogen bond with NH of residue
i+3 or 4th residue
Fig.3.42: Biochemistry 7th edition by Berg, Tymoczko and
Stryer
C O
N
H

24. Cont--
• Gly and Pro residues occurs in β-turn, Gly because its small and flexible
but Pro because peptide bond involves imino nitrogen assume cis conf,
a form a tight turn
• β-turn found near surface of protein where peptide groups of central two
aa residues in turn can hydrogen bond with water.

25. Cont--
 Loops are responsible for chain reversals
and overall shape.
 Turns and loops invariably lie on surfaces
of proteins and participate in recognition
role of proteins, such as recognition of
specific antigens by antibodies.
 Ex. part of antibody molecule has surface
loops (shown in red) that mediate
interactions with other molecules.
Fig.3.43: Biochemistry 7th edition by Berg, Tymoczko and Stryer

26. Tertiary Structure
 Overall 3-D arrangement of all atoms in protein is
referred as protein’s tertiary structure
 Myoglobin, oxygen carrier in muscle. It functions
both to store oxygen and to facilitate oxygen
diffusion in rapidly contracting muscle tissue.
 Capacity of myoglobin to bind oxygen depends on
presence of heme, a non-polypeptide prosthetic
group consisting of protoporphyrin IX and a central
iron atom.
Fig.3.44 b: Biochemistry 7th edition by Berg, Tymoczko and Stryer

27. Cont--
 Interior consists of nonpolar residues such as leucine, valine,
methionine, and phenylalanine
 Only polar residues inside are two histidine residues, play critical roles
in binding iron and oxygen.
 Outside of myoglobin, consists of both polar and nonpolar residues.

28. Higher level of structure
Useful to designate two major groups into fibrous and globular proteins:
 Fibrous Proteins with polypeptide chains arranged in long stands or
sheets
• Consist largely of a single type of 2⁰ structure and their 3⁰ structure is
simple
• Its structure provide support, shape and external protection
• Tertiary structure of the fibrous proteins: α-keratin, collagen and silk
fibroin

29. 1. α-keratin helix is a right handed α-helix,
arranged as a coiled coil
• These combine in higher-order structures
called protofilaments and protofibrils.
• Four protofibrils—32 strands of α-keratin
altogether—combine to form an
intermediate filament
• 2 strands of α-keratin, oriented in parallel
wrapped each other to form super-twisted
coiled coil
Fig.4.11: Biochemistry 7th edition by Berg, Tymoczko and Stryer

30. Cont--
• Coiled coil of this type are common structural elements in filamentous
proteins and in the muscle protein myosin
• Contribute to cell cytoskeleton (internal scaffolding in a cell)

31. Cont--
2. Collagen is main fibrous component of skin, bone,
tendon, cartilage, and teeth.
• Super helical twisting is right handed, opposite in sense
to left handed helix of α-chains.
• Three aa residue per turn. Glycine appears at every third
residue in a sequence.
• Hydrogen bonds within a strand are absent.
Fig.2.40: Biochemistry 7th edition by Berg, Tymoczko and Stryer
Fig.4.12 (a). Lehninger Principles of Biochemistry, 6th Ed.

32. Cont--
• Helix is stabilized by steric repulsion of pyrrolidine rings of proline and
hydroxyproline residues
• Seq of aa in collagen is repeating tripeptide unit, Gly-X(pro)-Y(4-Hyp)
• Only Gly residues accommodated at very tight junction bet individual α-
chains
• Pro and 4-Hyp residues permit sharp twisting of collagen helix
• Tight wrapping of α-chains in collagen triple helix provides tensile
strength

33. Cont--
Osteogenesis imperfecta: Abnormal bone formation in babies, at least
eight variants, with different degrees of severity.
• Results from substitution of an aa residue with larger R group (Cys or
Ser) for single Gly residue in α-chain in one or another collagen protein
• They disrupt Gly-X(pro)-Y(4-Hyp) repeat gives collagen its unique
helical structure

34. Cont--
 Globular Proteins: Different segments of polypeptide chain fold back
on each other, generates more compact shape
• Consists of variety of tertiary structure
• 3-D structure of typical globular protein consider assemblage of
polypeptide segments in α-helical and β conformations linked by
connecting segments
• Most of enzymes, transport proteins, motor proteins, regulatory
proteins, Ig are globular proteins

35. Quaternary Structure
🠶Some proteins contain two or more separate polypeptide chains or
subunits, may be identical or different.
🠶 Arrangement of these protein subunits in 3-D complexes constitutes
quaternary structure
🠶 Many multisubunit proteins have regulatory roles. Ex. each ribosome,
incorporates dozens of protein subunits along with a number of RNA
molecules
🠶 Quaternary structure implies non-covalent interaction that stabilise folded
polypeptides leads to multisubunit proteins.

36. Cont--
🠶 Multisubunit proteins/multimer have a number of identical (homomeric)
subunits (in symmetric arrangements and simplest) or non-identical
(heteromeric) subunits (asymmetric and complicated)
🠶 Repeating structural unit in multimeric protein whether a single or
groups of subunit is called protomer
🠶 Multimer with just a few subunits is called oligomer. First oligomeric
protein to have its 3D structure determined was Hemoglobin (Hb).

37. Cont--
• Ex. Hb (oxygen carrying protein) in
blood contains four polypeptide chains
and four heme prosthetic groups, in
which iron atoms are in ferrous (Fe2+)
state.
• Protein portion, called globin, consists
of two α-chains (141 residues) and two
β-chains (146 residues)
Fig.3.49: Biochemistry 7th edition by Berg, Tymoczko and Stryer

38. Cont--
🠶 Subunits of Hb are arranged in symmetric pairs, each pair having one
α and one β subunit.
🠶 Hb exists as an α2β2 tetramer or dimer of αβ protomers.
🠶 Subtle changes in arrangement of subunits within Hb molecule allow
it to carry oxygen from lungs to tissues with great efficiency.

39. Summary
🠶Most important element of primary structure is sequence of amino
acid residues.
🠶Nature of covalent bonds in polypeptide backbone places
constraints on structure.
🠶 Peptide bond has a partial double bond character that keeps entire
six-atom peptide group in a rigid planar configuration.
🠶N-Cα and Cα-C bonds can rotate to assume bond angles of φ and ψ,
respectively.

40. Cont--
🠶Gene-encoded primary structure of a polypeptide is sequence of its
amino acids.
🠶Secondary structure refers to stable arrangements of amino acid
residues giving rise to recurring structural patterns.
🠶 Folding of polypeptides into hydrogen-bonded motifs such as the α helix,
β-pleated sheet, β bends, and loops.

41. Cont--
🠶Tertiary structure is complete three-dimensional structure of a
polypeptide chain.
🠶When a protein has two or more polypeptide subunits, their
arrangement in space is referred to as quaternary structure

42. Reference Books
1) Harper’s Illustrated Biochemistry-30th edition
2) Biochemistry. 4th edition. Donald Voet and Judith G. Voet.
3) Biochemistry 7th edition by Jeremy M. Berg, John L. Tymoczko and Lubert
Stryer
4) Lehninger Principles of Biochemistry, 6th Ed by David L. Nelson, Michael
M. Cox
42

43. Thank You
