Protein structures determine their functions. There are four levels of protein structure: 1) Primary structure is the amino acid sequence 2) Secondary structure involves local patterns like alpha helices and beta sheets 3) Tertiary structure describes the overall 3D shape formed by secondary structures 4) Quaternary structure refers to the arrangement of multiple polypeptide chains The most common secondary structures are alpha helices, stabilized by hydrogen bonds between amino acids i and i+4, and beta sheets formed by hydrogen bonding between strands. Protein structure enables functions like catalysis, transport, and information transfer.

1. Protein Structure
RAFEEQ.C.M
Department of Biosciences

2. 2
Protein Functions
• Comes from Greek Work Proteios – PRIMARY
• Fundamental to virtually all cellular processes
– WHAT DON’T THEY DO!
– Proteins play a key role in biological processes. They can fulfill a vast
variety of tasks.
1. Enzymes/Cellular Signaling
 Enzymes catalyze the complex set of chemical reactions that are
collectively referred to as life.
 These chemical reactions are regulated by proteins, which act either
directly as components of enzymes or indirectly in the form of chemical
messengers, or their receptors.

3. 2. Antibodies
• The proteins of the immune system, such as the immunoglobulins, form
an essential biological defense system in higher animals.
3. Transport of materials
• Proteins are engaged in the transport and storage of biologically
important substances such as metal ions, oxygen, glucose, lipids, and
many other molecules.
“All of this results from the peculiar structure of proteins”
• Protein function can be understood only in terms of proteins structure. That
means the structure of a protein determines its biochemical function.

4. Protein Functions
• Three examples of protein functions
– Catalysis:
Almost all chemical reactions in a living
cell are catalyzed by protein enzymes.
– Transport:
Some proteins transports various
substances, such as oxygen, ions, and so
on.
– Information transfer:
For example, hormones.
Alcohol
dehydrogenase
oxidizes alcohols
to aldehydes or
ketones
Haemoglobin
carries oxygen
Insulin controls
the amount of
sugar in the
blood

5. Protein structure
• Primary structure is the linear sequence of amino acids in the
polypeptide chain(s) of a protein.
• Secondary structure- the local spatial arrangement of a
polypeptides backbone atoms without regard to the
conformations of its side chains.
Secondary Structure consists of local regions of poly peptide chains
that have a regular conformation (- helices, - sheets etc) which
is stabilized by H-bonds.

6. • Tertiary structure - the overall arrangement of secondary
structure elements.
• Tertiary Structure refers to the 3-D configuration of an entire
polypeptide chain .This includes - helices & - sheets and regions
that are globular or spherical
• Quaternary structure- the arrangement of several polypeptide
chains
• Quaternary Structure consists of number of polypeptide chains or
subunits joined by non covalent interactions.

8. Typical bonds in protein molecules
Covalent bonds
Non-covalent bonds
Peptide bond
Disulfide bond
Hydrogen bond
Ionic bond
Hydrophobic interactions

9. Primary Structure
It’s the sequence of amino acids linked through peptide bonds, Covalent
backbone of the protein.
H2N-Glu-Ala-Val-Ser-Leu-Ala-Lys-Cys-COOH

10. • The particular sequence of amino acids in a peptide or protein is referred to as
the primary structure.
--For example, a hormone that stimulates the thyroid to release thyroxine (TSH)
consists of a tri-peptide Glu-His-Pro.
--Although other sequences are possible for these three amino acids, only the
tri-peptide with the Glu-His-Pro sequence of amino acids has hormonal activity.
-- Thus the biological function of peptides as well as proteins depends on the
order of the amino acids.

11. Amino Acids
• While their name implies that amino acids are compounds that contain an
—NH2 group and a —COOH group, these groups are actually present as —
NH3
+ and —COO– respectively.
• More than 700 amino acids occur naturally, but 20 of them are especially
important.
• These 20 amino acids are the building blocks of proteins. All are -amino
acids.
• They differ in respect to the
group attached to the  carbon.

14. 20 Amino acids
Glycine (G)
Glutamic acid (E)
Asparatic acid (D)
Methionine (M)
Threonine (T)
Serine (S)
Glutamine (Q)
Asparagine (N)
Tryptophan (W)
Phenylalanine (F)
Cysteine (C)
Proline (P)
Leucine (L)
Isoleucine (I)
Valine (V)
Alanine (A)
Histidine (H)
Lysine (K)
Tyrosine (Y)
Arginine (R)
White: Hydrophobic, Green: Hydrophilic, Red: Acidic, Blue: Basic

15. 15
Amino acids exist as a zwitterion:
a dipolar ion having both a formal positive and formal negative
charge (overall charge neutral).
Amino acids are amphoteric:
They can react as either an acid or a base. Ammonium ion
acts as an acid, the carboxylate as a base.
Isoelectric point (pI):
The pH at which the amino acid exists largely in a neutral,
zwitterionic form (influenced by the nature of the side chain)

16. 16
Peptide bonds
• Amino Acids connect via PEPTIDE BOND

19. Peptide Bond
• Joins amino acids
• Covalent, strong bond
• Not broken by usual denaturing agents like heating or
high salt concentration
• Can be broken by Prolonged exposure to strong acids
or base at elevated temperatures or by enzyme
digestion.
• Rigid and planar
• 40% double bond character
– Caused by resonance

20. – Results in
shorter bond
length
Peptide Bond Lengths

21. – Partial double bond character (distance is 1.33 Å (angstroms)
which is midway in a single bond 1.45 Å and a double bond
1.23Å)
– Double bond disallows rotation around the bond between
Carbonyl carbon and the nitrogen of the peptide bond
– The bonds between the alpha carbon and the alpha amino
groups can be freely rotated.
– But they are limited by the size and character of the R Groups

22. Secondary structure

23. 23
• Backbone can swivel:
DIHEDRAL ANGLES
• 2 per Amino Acid
• Proteins can be 100’s of
Amino Acids in length!
– Lots of freedom of
movement

25. Many of the possible conformations about an α-carbon between
two peptide planes are forbidden because of steric crowding.
Several noteworthy examples are shown here.

26. Backbone Torsion Angles

27. Backbone Torsion Angles
• ω angle tends to be planar (0º - cis, or 180 º - trans) due to
resonance stabilization
• φ and ψ are flexible, therefore rotation occurs here
• However, φ and ψ of a given amino acid residue are limited due
to steric hindrance
• Only 10% of the area of the {φ, ψ} space is generally observed
for proteins
• First noticed by G.N. Ramachandran

28. G.N. Ramachandran
8 October 1922 – 7 April 2001
• Indian Physicist, Student of CV Raman.
• Used computer models of small polypeptides
to systematically vary φ and ψ with the objective
of finding stable conformations
• For each conformation, the structure was
examined for close contacts between atoms
• Atoms were treated as hard spheres with
dimensions corresponding to their van der Waals
radii

29. Ramachandran Plot
• Plot of φ vs. ψ
• Repeating values of φ and ψ along the chain result in regular
structure
• By Applying the values of φ and ψ in the Ramachandran plot we can
predict the secondary structure of a protein
• For example, repeating values of φ ~ -57° and ψ ~ -47° give a right-
handed helical fold (the alpha-helix)

30. • Therefore, φ and ψ angles which cause spheres to collide correspond to sterically
disallowed conformations of the polypeptide backbone

32. Ramachandran Plot
• White = sterically disallowed conformations (atoms in the
polypeptide come closer than the sum of their van der Waals radii)
• Red = sterically allowed regions (namely right-handed alpha helix
and beta sheet)
• Yellow = sterically allowed if shorter radii are used (i.e. atoms
allowed closer together; brings out left-handed helix)

33. • The structure of cytochrome C-256 shows many segments of helix and the
Ramachandran plot shows a tight grouping of φ, ψ angles near -50,-50
alpha-helix
cytochrome C-256 Ramachandran plot

34. • Linus Pauling and Robert Corey describes the -helix in 1951
• In this structure the polypeptide backbone is tightly wound around an
imaginary axis drawn longitudinally through the middle of the helix in a
right-handed manner.
• R groups of the amino acid residues protrude outward from the helical
backbone.
• It helps to avoid the steric interference with the polypeptide backbone
• The repeating unit is a single turn of the helix, having a pitch of 5.4 Å.
• Each helical turn includes 3.6 amino acid residues.
-HELIX

35. • The  helices of proteins have an
average length of 12 residues,
which corresponds to over three
helical turns, and a length of 18 Å.
• Generally, about one-fourth of all
amino acid residues in
polypeptides are found in helices,
the exact fraction varying greatly
from one protein to the next.

36. • Why does the helix form more readily than many other possible conformations?
• The structure is stabilized by a hydrogen bond
• A helix makes optimal use of internal hydrogen bonds.
• Hydrogen bonding is seen between the nth and n+4 the residue
• This results in a strong hydrogen bond has the nearly optimum distance of 2.8 Å
• Within the helix, every peptide bond (except those close to each end of the helix)
participates in such hydrogen bonding.
• Each successive turn of the helix is held to adjacent turns by three to four hydrogen
bonds.
• All the hydrogen bonds combined give the entire helical structure considerable
stability.

37. The α-Helix
Four different representations of the α-helix
First proposed by Linus Pauling and Robert Corey in 1951
A ubiquitous component of proteins, stabilized by H bonds

38. • Residues per turn: 3.6
• Rise per residue: 1.5 Angstroms
• Rise per turn (pitch): 3.6 x 1.5A = 5.4 Angstroms
• The backbone loop that is closed by any H-bond
in an alpha helix contains 13 atoms
• phi = -57 degrees, psi = -47 degrees
• The non-integral number of residues per turn was
a surprise to crystallographers

39. Amino Acid Sequence Affects Helix Stability;
• Not all polypeptides can form a stable helix.
• Interactions between amino acid side chains can stabilize or destabilize this
structure.
• For example, if a polypeptide chain has a long block of Glu residues, this
segment of the chain will not form a helix at pH 7.0.
• The negatively charged carboxyl groups of adjacent Glu residues repel each
other so strongly that they prevent formation of the helix.
• Another constraint on the formation of the helix is the presence of Pro or Gly
residues.

40. • In proline, the nitrogen atom is part of a rigid ring and rotation about the NOC
bond is not possible. Thus, a Pro residue introduces a destabilizing kink in an
helix.
• In addition, the nitrogen atom of a Pro residue in peptide linkage has no
substituent hydrogen to participate in hydrogen bonds with other residues.
• For these reasons, proline is only rarely found within an helix.
• Glycine occurs infrequently in helices for a different reason:
• It has more conformational flexibility than the other amino acid residues.
Polymers of glycine tend to take up coiled structures quite different from an
helix.

41. Five different kinds of constraints affect the stability of a helix:
• (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 Pro and Gly residues, and
• (5) The interaction between amino acid residues at the ends of the helical
segment and the electric dipole inherent to the helix

42. Other (Rarer) Helix Types - 310
• Less favorable geometry
• Pitch is 6.0Å
• Rise per residue is 2.0 Å
• 3 residues per turn with n+3 not n+4
• Hence narrower and more elongated
• Usually seen at the end of an alpha helix
• frequently occurs as a single turn transition between the end
of an a-helix and the next portion of the polypeptide chain

43. Other (Very Rare) Helix Types - Π
• Less favorable geometry
• p helix (4.416 helix): p=5.2Å
• Rise per residue= 1.3 Å
• 4 residues per turn with i+5 not i+4
• Squat and constrained

44. The Beta-Pleated Sheet
• Also first postulated by Pauling and Corey, 1951
• Formed through by side-by-side alignment of polypeptide strands
• Polypeptide chains are held together by hydrogen bonds between the
peptide chains.
• R groups of extend above and below the sheet
• Rise per residue:
– 3.47 Angstroms for antiparallel strands
– 3.25 Angstroms for parallel strands
– Each strand of a beta sheet may be pictured as a helix with two
residues per turn

45. An antiparallel β-pleated sheet. R groups project alternately above and below the plane of the
sheet. Sheet structure is derived from the tetrahedral placement of substituents on the α
carbon atoms. This is the more stable form of a β-sheet.

46. • Comparison of β-sheet with α-helix
• The beta pleated sheet differs markedly from the a helix in that it is a
sheet rather than a rod.
• The polypeptide chain in the beta pleated sheet is almost fully extended
rather than being tightly coiled as in the a helix.
• The axial distance between adjacent amino acids is 3.5 A, in contrast with
1.5 A for the a helix.
• beta pleated sheet is stabilized by hydrogen bonds between NH and CO
groups in different polypeptide strands, whereas in the a helix the
hydrogen bonds are between NH and CO groups in the same polypeptide
chain.

47. • Adjacent strands in a beta pleated sheet can run in the
same direction (parallel beta sheet) or in opposite
directions (antiparallel beta sheet).
• H-bonds perpendicular to long axis
• β-sheets are composed of two or more polypeptide
chains or extended segments of the same polypeptide

48. TYPES OF -PLEATED SHEET
Antiparallel Parallel
Antiparallel is more stable than parallel
Both models are found in proteins

50. β - turns
Beta-turn loops allow for protein
compaction, since the hydrophobic amino
acids tend to be in the interior of the
protein, while the hydrophilic residues
interact with the aqueous environment.
(aka beta bend, tight turn)
Permits the change of direction of the peptide chain
to get a folded structure.
carbonyl C of one residue is H-bonded to the amide
proton of a residue three residues away
proline and glycine are prevalent in beta turns

51. 51
Other Secondary Structures – Loop or Coil
• Often functionally significant
• Different types
– Hairpin loops (aka reverse turns) – often between
anti-parallel beta strands
– Omega loops – beginning and end close (6-16
residues)
– Extended loops – more than 16 residues

52.  Complete three-dimensional shape of
a given protein.
 Represent the spatial relationship of
the different secondary structures to
one another within a polypeptide
chain and how these secondary
structures themselves fold into the
three-dimensional form of the
protein.
Tertiary structure
The spiral regions represent sections of the
polypeptide chain that have an α-helical
structure, while the broad arrows represent β-
pleated sheet structures.

53.  A domain is a basic structural unit within a
protein molecule.
 Part of protein that can fold into a stable
structure independently.
 Different domains can possess different
functions.
 Proteins can have one to many domains
depending on protein size.
 A polypeptide with 200 amino acids consists
of two or more domains.
 Domains are usually connected with
relatively flexible areas of protein.
Protein domain
Pyruvate kinase (a
monomeric protein):
three domains
Tertiary structure: Describes the relationship of different
domains to one another within a protein.

54. Tertiary structure is based on various types of interactions between
the side-chains of the peptide chain.

55. STABILIZING INTERACTIONS OF TERTIARY STRUCTURES

56. Globular Proteins
 Globular proteins fold up into compact, spherical
shapes.
 Their functions are related to cell metabolism:
biosynthesis and biodegradation, transport, catalytic
function.
 Hydrophobic R-groups are oriented into inner part of
the protein molecule, while hydrophilic R-groups are
pointed towards molecule edges.
 Globular proteins are water soluble.

57. Example: myoglobin
Globular protein that stores
oxygen in muscles
A single peptide chain that
is mostly -helix
O2 binding pocket is formed
by a heme group and specific
amino acid side-chains that
are brought into position by
the tertiary structure

58.  Much or most of the polypeptide chain is
parallel to a single axis
 Fibrous proteins are often mechanically
strong and highly cross-linked
 Fibrous proteins are usually insoluble
 Usually play a structural role
Fibrous proteins

59. Fibrous Proteins: Keratins
• For example, -keratins are fibrous proteins that make
hair, fur, nails and skin
- hair is made of twined fibrils
- the -helices are held together by disulfide bonds

60. Fibrous proteins: Fibroin
Fibroin
 Fibroins are the silk proteins. They also form the spider webs
 Made with a -sheet structures with Gly on one face and
Ala/Ser on the other
 Fibroins contain repeats of [Gly-Ala-Gly-Ala-Gly-Ser-Gly-Ala-
Ala-Gly-(Ser-Gly-Ala-Gly-Ala-Gly)8]
 The -sheet structures stack on top of each other
 Bulky regions with valine and tyrosine interrupt the -sheet
and allow the stretchiness

61. Collagen is formed from tropocollagen subunits. The
triple helix in tropocollagen is highly extended and
strong.
Features:
 Three separate polypeptide chains arranged as a
left-handed helix (note that an a-helix is right-
handed).
 3.3 residues per turn
 Each chain forms hydrogen bonds with the other
two: STRENGTH!
 Nearly one residue out of three is Gly
 Proline content is unusually high
 Many modified amino acids present:
 4-hydroxyproline
 3-hydroxyproline
 5-hydroxylysine
 Pro and HydroxyPro together make 30% of amino
acids.
Collagen amino
acid composition:
Fibrous proteins: Collagen is a Triple Helix

62. Covalent cross-links in collagen: alteration of
mechanical properties of collagen
Catalyzed by lysyl amino oxidase

63. 1. Compact protein structure Extended protein structure
2. Soluble in water (or in lipid Insoluble in water (or in lipid
bilayers) bilayers)
3. Secondary structure is а complex Secondary structure is simple
with a mixture of a-helix, b-sheet with predominant one type only
and loop structures
4. Quaternary structure is held Quaternary structure is usually
together by noncovalent forces held together by covalent
bridges
5. Functions in all aspects of Functions in structure of the
body metabolism (enzymes, transport, or cell (tendons, bones, muscle,
immune protection, hormones, etc). ligaments, hair, skin)
Globular proteins vs Fibrous proteins

64. Quaternary structure of proteins
Monomeric proteins:
– built of a single polypeptide chain.
Oligomeric proteins:
– built of more than one polypeptide chains called
subunits or monomers.

65.  Quaternary structure describes the joining of two
or more polypeptide subunits.
 The subunits each have their own tertiary
structure.
 Bonds – non-covalent interactions.
 Subunits can either function independently or
work co-operatively.
 Dissociation of a subunit results in loss of function.

66. For example: Hemoglobin
 A globular protein that consists of four subunits (2α and 2β, of two
different types (α and β)
 Each subunit contains a heme group for O2 binding
 Binding O2 to one heme facilitates O2 binding by other subunits
 Replacement of even one amino acid in primary structure with
another amino acid is critical for the function of the protein.

67. Structural organization of proteins: Summary