Protein chemistry, Biochemistry

1. PROTEIN CHEMISTRY-III
By Dr ANURAG YADAV

2. CONTENT:
• Globular hemeproteins.
-myoglobin
-hemoglobin
• Structure of collagen.
• Study of the Protein structure.

3. Globular Hemeproteins
• Group of specialized proteins that contain heme as tightly
bound prosthetic group.
• In the hemoglobin and myoglobin, heme group serve to
reversibly bind oxygen.

4. A. Structure of heme:
- complex of Protoporphyrin IX & ferrous iron.
- The iron is held in center of heme molecule by bonds to 4
Nitrogen of porphyrin ring.
- Heme ferrous can form another 2 bonds.
- In hemoglobin & myoglobin one of these position is
coordinated to side chain of histidine residue of globin
molecule.

5. Structure of Myoglobin :
1. α -helical content:
- Compact molecule with 80% of α-
helical polypeptide.
- Folded into 8 stretches.
- Labeled from A-H.
- Terminated either by Proline/ β-
bend/ loops stabilized by Hydrogen
bonds & ionic bonds.

6. 2.Location of Polar & Non-Polar
groups:
3. Binding of heme:
- The heme group of myoglobin sits in
a crevice in the molecule
- Lined by the non-polar amino acids.
- The proximal histidine F8 binds
directly to iron , but the distal E7,
helps to stabilize the binding of
oxygen to ferrous iron.

7. Structure of Hemoglobin:

8. 1. Quaternary structure of hemoglobin:
- composed of two identical dimers (αβ)1 & (αβ)2.
- Polypeptide chain within each dimer held together primarily
by the hydrophobic interaction.
- Interchain hydrophobic interaction form strong association
btwn α-subunit & β-subunit in the dimer.
- Ionic & hydrogen bonds also occur btwn members of dimer.

10. - The two dimers are able to move with respect to each other
- Held primarily by the polar bonds.
- Weaker interactions btwn these mobile dimers.
- The binding of the O2 to heme iron pulls the iron into the
plane of the heme.

11. It is of two forms:
a. T form:
- deoxy form
- the 2 αβ dimers interact through
network of ionic bonds &
hydrogen bonds that constrain
the movement of polypeptide
chains.
- it is low Oxygen-affinity form
b. R form:
- oxy form
- binding of O2 cause rupture of
some of the ionic bonds &
hydrogen bonds btwn the
dimers.
- In which the chains have more
freedom of movement.
- it is an high Oxygen-affinity form.

13. STRUCTURE OF COLLAGEN
1. AMINO ACID Sequence:
- Rich in Proline and Glycine ( imp in
formation of triple helix).
- Proline facilitate formation of helical
conformation of each α chain of helix
(because its ring structure cause kink in
the peptide chain).
- Glycine ( smallest aa) found every third
position.
- It is repetitive part with sequence as -
gly-X-Y- (X frequently is Proline, Y often is
hydroxyproline).

14. 2. Triple helix formation
3. Hydroxyproline and hydroxylycine:
- These are seen as part of collagen
- Result from hydroxylation of some
Proline& lysine after incorporation
into polypeptide chain.
- Imp in stabilizing the triple helix (
maximizes the Interchain hydrogen
bonding).

15. Collagen diseases:
- Ehlers-Danlos syndrome
- Osteogenesis imperfecta

16. Study of Protein Structure:
• The first protein to be sequenced was insulin by Sanger.
• Before studying the structure, first pure sample of the protein
has to be available.
• Various proteins are extracted and purified by-
Chormatography technique
• Purity is studied by the Electrophoresis.
• Molecular weight by the mass spectroscopy.
-Ion exchange
-Adsorption
-Partition
-Paper chromatography
-Affinity
-HPLC

17. Steps to Determining the Primary structure:
- Determining the number of polypeptide chains in protein.
- Determine amino acid composition of polypeptide chain by
complete hydrolysis.
- Identification if N- & C- terminals
- Site specific enzymes used for hydrolysis of chain into
peptides.
- Separation and purification of each peptides and then
analyzing amino acid sequence.

18. Dansyl chlorideFDNB
Steps in Edman's degradation process

19. • Finger printing Method( Ingram’s Technique)
• Automatic sequencing
• study of Higher levels of Proteins structure.
- X-ray diffraction.
- nuclear magnetic resonance(NMR).