- Proteins are composed of amino acid polymers called polypeptides linked by peptide bonds. They form complex 3D structures through folding at various levels - primary, secondary, tertiary, and quaternary. - Secondary structure includes alpha helices, beta sheets, and beta turns formed by hydrogen bonding between amino acids in the polypeptide chain. - Tertiary structure is the overall 3D shape of a protein formed by interactions between amino acid side chains. Quaternary structure involves the assembly of multiple protein subunits. - Myoglobin and hemoglobin are oxygen-binding proteins with heme prosthetic groups. Hemoglobin has a cooperative binding mechanism that allows for oxygen delivery and release in tissues.

1. Fundamentals of Protein
Structure
The Three Dimensional
Structure of Proteins

2. • Proteins are major components of all
cellular systems
• Proteins consist of one or more
linear polymers called polypeptides
• Proteins are linear and never
branched
• Different AA’s are linked together
via PEPTIDE bonds
• The individual amino acids within a
protein are known as RESIDUES
• The smallest known Peptide is just
nine residues long - oxytocin
• The largest is over 25,000 residues -
the structural protein titin
Introduction

3. Classification of proteins
A. Functional Basis
Nothing can compare with the versatility of proteins. Their functionality and usage
in organisms is unrivalled.
• Enzymes: Catalytic activity Carbonic Anhydrase, Lipases
• Structural: Provide support collagen fibers, elastin, keratin
• Contractile Proteins: Actin and Mayosin
• Storage Proteins: Ovalbumin, ferritin, Casein
• Blood clotting: Fibrinogen
• Cytoskeleton: Tubulin
• Hormones: Insulin and Glucagon
• Transportation: bind and carry ligand molecules. Hemoglobin
• Defense Mechanism: Antibodies

4. B. Structural Basis :
Globular: Complex folds, irregularly shaped tertiary structures
Fibrous: Extended, simple folds -- generally structural proteins
C. Cellular localization definition:
Membrane: In direct physical contact with a membrane;
generally water insoluble.
Soluble: Water soluble; can be anywhere in the cell.
D. Structural Composition:
Non Conjugated: No prosthetic Group
Conjugated: Proteins associated with organic or non organic
non proteinous part
• Glycoproteins
• Metalloproteins
• Phophoproteins
• Lipoproteins
• flavoproteins

5. Levels of Protein Structure

6. Primary structure = order of amino
acids in the protein chain

7. Amino Acids Are Joined By Peptide
Bonds In Peptides
- a-carboxyl of one amino acid is joined to a-amino of a
second amino acid (with removal of water)
- only a-carboxyl and a-amino groups are used, not R-
group carboxyl or amino groups

8. Properties of Peptide Bond
•Bond Length: 1.32A (Double bond:1.21A, Single bond: 1.47A) due
to resonance or partial sharing of electrons.
* Resonance: state of adjustment that produces resonance in a system
•Planar: On one side of the Plane.
•Rigid: not able to move or orient itself (due to planner
Hence, it posses 40% double bond characters

9. Conformation of Polypeptide Chain is defined by:
2 Torsion Angle/Dihedral angles: Angles between two planes
1.Φ (phi): rotation around alpha carbon and nitrogen
2.Ψ (psi): rotation around alpha carbon and carbon
•Ψ and φ angles will cause the hydrogen and oxygen and other
residue to collide
•Many chain rotations are restricted to Ψ and φ.
•This restricts the number of conformations in proteins

10. • Many angles of rotation are possible only a few are energetically
favorable
• By convention Ψ and φ are 180° (or –180°) when the first and
fourth atoms are farthest apart and the peptide is fully extended.
• In a protein, some of the conformations shown here (e.g., 0°)
are prohibited by steric overlap of atoms.

11. • Every amino acid has its own set of angles
defining direction of possible rotation
within the molecule.
Ramachandran plots
•Ramachandran plot shows the distribution of f and y dihedral angles that
are found in a protein
•Some Ψ and φ combinations are very unfavorable because of steric crowding of
backbone atoms with other atoms in the backbone or side-chains.
•Some Ψ and φ combinations are more favorable because of chance to form
favorable H-bonding interactions along the backbone (Blue-shaded areas).
•shows the common secondary structure elements reveals regions with unusual
backbone structure

12. Secondary Structure: = local folding of residues into regular patterns
• The chains of amino acids fold or turn upon themselves
• Held together by hydrogen bonds between (non-adjacent) amine
(N-H) and carboxylic (C-O) groups
• H-bonds provide a level of structural stability Secondary structure
• For example Fibrous proteins
3 Major Types
• α-Helix
• β-Conformation or Sheets
• β-Turns

13. The a-helix • In the a-helix, the carbonyl oxygen of
residue “i” forms a hydrogen bond with
the amide of residue “i+4”.
• Although each hydrogen bond is
relatively weak in isolation, the sum of
the hydrogen bonds in a helix makes it
quite stable.
• The propensity of a peptide for forming
an a-helix also depends on its sequence.
• Right-handed helix with 3.6 residues (5.4
Å) per turn
• Peptide bonds are aligned roughly parallel
with the helical axis
• Side chains point out and are roughly
perpendicular with the helical axis

14. • Not all polypeptide sequences adopt a helical structures
• Small hydrophobic residues such as Ala and Leu are strong helix formers
• Pro acts as a helix breaker because the rotation around the N-Ca bond is
impossible
• Gly acts as a helix breaker because the tiny R group supports other conformations
ΔΔG is the difference in free-energy change relative to that for alanine

15. The b-sheet
• The backbone is more extended, the planarity of the peptide
bond and tetrahedral geometry of the a-carbon create a pleated
sheetlike structure
• Sheet-like arrangement of backbone is held together by
hydrogen bonds between the more distal backbone amides and
carbonyl oxygen.
• Side chains protrude from the sheet alternating in up and down
direction

16. • Core of many proteins is the b sheet
• Parallel or antiparallel orientation
of two chains within a sheet are
possible
• In parallel b sheets the H-bonded
strands run in the same direction
• In antiparallel b sheets the H-bonded
strands run in opposite directions
Beta strand is an extended structure
3.5A between R groups in sheet
compared to 1.5 in alpha helix
• The propensity of a peptide for
forming b-sheet also depends on its
sequence.
• Most ß strands in proteins are 5 to 8
AA long.

17. b turns
 b -turns occur frequently whenever strands in b sheets change the direction
• The 180° turn is accomplished over four amino acids
• The turn is stabilized by a hydrogen bond from a carbonyl oxygen to amide
proton three residues down the sequence.
• ß Turns consist of 3-4 amino acids that form tight bends.
• Glycine and proline are common in turns. Longer connecting segments
between ß strands are called loops.
• Proline in position 2 or glycine in position 3 are common in b-turns
b-turns allow the protein backbone to make abrupt turns.
• Again, the propensity of a peptide for forming b-turns depends on its
sequence.

18. Protein Structure
Individual PROTEINS’ domains may and
generally do consist of a combination of
secondary structures

19. Tertiary structure = global folding of a protein chain
• The polypeptide folds and coils to form a complex 3D shape
• Caused by interactions between R groups (H-bonds, disulphide bridges, ionic
bonds and hydrophilic / hydrophobic interactions)
• Tertiary structure may be important for the function (e.g. specificity of active
site in enzymes)

20. Proteins are commonly described as either being fibrous or
globular in nature.
1. Fibrous proteins: typically insoluble; made from a single
secondary structure
2. Globular proteins: water-soluble globular proteins, multiple
secondary structure are involved

21. Fibrous proteins
• The fundamental structural unit is a simple
repeating element of secondary structure.
• All fibrous proteins are insoluble in water, a
property conferred by a high concentration of
hydrophobic amino acid residues both in the
interior of the protein and on its surface.
• These hydrophobic surfaces are largely
buried as many similar polypeptide chains
are packed together to form elaborate
supramolecular complexes.

22. Globular proteins
Protein Folding
• Non-covalent bonds within and between Peptide chains are
as important in their overall conformation and function
• Weak non-covalent interactions including IONIC,
HYDROGEN, and other HYDRPHOBIC INTERACTIONS
will hold the protein in its functional shape
• Disulfide bonds (S-S) form between adjacent -SH groups on
the amino acid cysteine AND these Cross linkages can be
between 2 parts of a protein or between 2 subunits
• The peptide bond allows for rotation around it and therefore the protein can fold
and orient the R groups in favorable positions

23. Motif, also called a super secondary
structure or fold. A motif is simply a
recognizable folding pattern involving two
or more elements of secondary structure
and the connection(s) between them.
It is a folding pattern that can describe a
small part of a protein or an entire
polypeptide chain
Domain
A domain, is a part of a polypeptide chain that
is independently stable or could undergo
movements as a single entity with respect to the
entire protein.
Polypeptides with more than a few hundred
amino acid residues often fold into two or more
domains, sometimes with different functions

24. Quaternary structure = Higher-order assembly of proteins
• Quaternary structure is formed by spontaneous assembly of individual
polypeptides into a larger functional cluster
• Oligomeric Subunits (protomers) are arranged in Symmetric Patterns
* protomer is the structural unit of an oligomeric protein.
• The interaction between multiple polypeptides or prosthetic groups
• A prosthetic group is an inorganic compound involved in a protein (e.g.
the heme group in haemoglobin)

25. Examples of other quaternary structures
Tetramer Hexamer Filament
SSB DNA helicase Recombinase
Allows coordinated Allows coordinated DNA binding Allows complete
DNA binding and ATP hydrolysis coverage of an
extended molecule

26. • Myoglobin is an iron- and oxygen-
binding protein found in the muscle
tissue .
• It is distantly related to hemoglobin
which is the iron- and oxygen-binding
protein in blood, specifically in the red
blood cells
• Å single subunit 153 amino acid residues
• 121 residues are in an a helix. Helices
are named A, B, C, …F. The heme
pocket is surrounded by E and F but not
B, C, G, also H is near the heme.
• Non-polar R-groups tend to be buried in
the cores of soluble proteins
Blue = non-polar R-group
Red = Heme
Myoglobin

27. Myoglobin facilitates rapidly respiring muscle tissue
➢ The rate of O2 diffusion from capillaries to tissue is slow because of the
solubility of oxygen.
➢ Myoglobin increases the solubility of oxygen, and also facilitates oxygen
diffusion.
➢ Oxygen transported over large distances by iron, incorporated into a protein-
bound prosthetic group called heme (or haem) having high oxygen carrying
capacity.
➢ Heme consists of a complex organic ring structure, protoporphyrin, to which is
bound a single iron atom in its ferrous (Fe'*) state.
➢ The iron atom has six coordination bonds, four to nitrogen atoms that are part
of the flat porphyrin ring system and two perpendicular to the porphyrin. The
coordinated nitrogen atoms (which have an electron-donating character) help
prevent conversion of the heme iron to the ferric (Fe3+) state.
➢ Iron in the ferrous (Fe2+) state binds oxygen reversibly; in the Fe3+ state it does
not bind oxygen. Heme is also found in many oxygen-transporting proteins, as
well as in some proteins, such as the cytochromes that participate in oxidation-
reduction (electron-transfer) reactions.
➢ Oxygen storage is also a function because Myoglobin concentrations are 10-
fold greater in whales and seals than in land mammals

28. Hemoglobin
• Different Subunit Proteins (heteromeric), 2 a globin subunits and 2 b globin subunits
• Composed of four subunits, each containing a heme group: a ring-like structure Porphyrin
with a central iron atom that binds oxygen.
• Extensive interactions between unlike subunits a2-b2 or a1-b1 interface has 35 residues
while a1-b2 and a2-b1 have 19 residue contact.
• a2,b2 dimer which are structurally similar to myoglobin
• Transports oxygen from lungs to tissues. O2 diffusion alone is too poor for transport in
larger animals.
• Solubility of O2 is low in plasma i.e. 10-4 M. But bound to hemoglobin, [O2] = 0.01 M or
that of air.
• Two alternative O2 transporters are;
Hemocyanin, a Cu containing protein (Found in Arthropods and Mollusca).
Hemoerythrin , a non-heme containing protein (marine invertebrate).

29. • Protoporphyrin binds oxygen to the sixth ligand of Fe(II) out of the plane of
the heme. The fifth ligand is a Histidine, F8 on the side across the heme
plane.
• His F8 binds to the proximal side and the oxygen binds to the distal side.
• The heme alone interacts with oxygen such that the Fe(II) becomes oxidized
to Fe(III) and no longer binds oxygen.
• The heme group is nonplanar when it is not bound to oxygen; the iron atom is
pulled out of the plane of the porphyrin, toward the histidine residue to which
it is attached.
• This nonplanar configuration is characteristic of the deoxygenated heme
group, and is commonly referred to as a "domed" shape.
• However, when the Fe in the heme group binds to an oxygen molecule, the
porphyrin ring adopts a planar configuration and hence the Fe lies in the plane
of the porphyrin ring
Function of the Hemoglobin

30. Mechanism of Oxygen binding to
hemoglobin
Tense “T” state
• Binds oxygen with low affinity
• Favoured at low oxygen concentration
Relaxed “R” State
• Binds oxygen with high affinity
• Binding energy with oxygen stabilizes R state
• Becomes predominant as oxygen concentration
increases

31. • As each O2 molecule binds, it alters the conformation of haemoglobin, making
subsequent binding easier (cooperative binding)
• This means haemoglobin will have a higher affinity for O2 in oxygen-rich areas (like the
lung), promoting oxygen loading
• Conversely, haemoglobin will have a lower affinity for O2 in oxygen-starved areas (like
muscles), promoting oxygen unloading
• The oxygen dissociation curve for adult haemoglobin is sigmoidal (i.e. S-shaped) due to
cooperative binding
• There is a low saturation of haemoglobin when oxygen levels are low (haemoglobin
releases O2 in hypoxic tissues)
• There is a high saturation of haemoglobin when oxygen levels are high (haemoglobin
binds O2 in oxygen-rich tissues)