1. CHMI 2227 - E.R. Gauthier, Ph.D. 1 CHMI 2227E Biochemistry I Proteins: - Quaternary structure

2. CHMI 2227 - E.R. Gauthier, Ph.D. 2 Quaternary structure

3. CHMI 2227 - E.R. Gauthier, Ph.D. 3 Quaternary structure Quaternary structure involves several polypeptides:  Oligomers  Heteromers These subunits interact with each other through the usual weak interaction forces (H bonds, Van der Waals, ionic interactions, hydrophobic interactions) and/or though disulfide bonds; For aquous proteins, frequently, but not always, the interface between two subunits is made of hydrophobic amino acids. For membrane-bound proteins, the amino acids at the interface between the subunits are usually hydrophilic; Porin: a trimeric membrane-bound protein hydrophobic hydrophilic

4. CHMI 2227 - E.R. Gauthier, Ph.D. 4 Quaternary structure Hemoglobin Made up of 4 polypeptide chains:  2 copies of  -subunit (or HbA): yellow and blue;  2 copies of  -subunit (or HbB): red and pink Each subunit binds its own heme group: so each subunit can bind O 2 Each subunit is highly similar in structure to myoglobin; Both hemoglobin and myoglobin bind O 2 in a very similar fashion

5. CHMI 2227 - E.R. Gauthier, Ph.D. 5 Quaternary structure Hemoglobin HbA vs myoglobin HbAMyoglobin

6. CHMI 2227 - E.R. Gauthier, Ph.D. 6 Quaternary structure Oxygen binding by Hb and myoglobin 4 major residues surround the heme group:  Phe 43  His 64  Val 68  His 93 These amino acids create a hydrophobic environment while help hold the heme group in place; Also: His 93 binds the Fe 2+ atom;

7. CHMI 2227 - E.R. Gauthier, Ph.D. 7 Quaternary structure Oxygen binding by Hb and myoHb O 2 binds the Fe 2+ atom of the heme group, and is held in place with His 64; Oxygen-bound myoglobin/Hb is called oxymyoglobin/oxyHb Oxygen-free myoglobin/Hb is called deoxymyoglobin/deoxyHb Now, if both Myo and Hb can bind O 2, why is it that Hb is a multimeric protein, while myoglobin is monomeric??? WHY????

8. CHMI 2227 - E.R. Gauthier, Ph.D. 8 Quaternary structure Oxygen binding by Hb and myoHb A B O 2 binding to myoglobin shows a simple equilibrium where the amount of O 2 bound-myoglobin (y) directly depends on the concentration of O 2 present; However, O2 binding to Hb is more complex:  At low O 2 concentration, very little Hb binds O 2 even as the concentration of O 2 increases (part A of the Hb curve);  However, at a certain threshold of O 2 concentration, Hb becomes rapidly saturated with O 2 (part B of the Hb curve);

9. CHMI 2227 - E.R. Gauthier, Ph.D. 9 Quaternary structure Oxygen binding by Hb and myoHb A B At high O 2 concentrations, both myoglobin and Hb are saturated, meaning there are no more O 2 -binding spots available. Interestingly: the affinity of myoglobin and Hb for oxygen varies by a factor of 10:  Only 2.8 Torr are required to get 50% of myoglobin saturated;  However, 26 Torr are required to half-saturate Hb.

10. CHMI 2227 - E.R. Gauthier, Ph.D. 10 Quaternary structure O 2 binding changes the 3-D shape of Hb In the deoxyHb form, Fe 2+ is bonded to 5 ligands: His 93 and 4 amines from the heme group; When one subunit of Hb binds O 2, the Fe 2+ atom moves foward the plane of the heme group, pulling with it the His 93 and the  -helix; This causes a slight but significant change in the tertiary structure of all the other Hb subunits, even if they are in the deoxyHb form;

11. CHMI 2227 - E.R. Gauthier, Ph.D. 11 Quaternary structure O 2 binding changes the 3-D shape of Hb The consequence of this slight change in conformation is an increase in the affinity of these other Hb subunits for O 2 ; This phenomenon, where a change in the shape in one subunit trigger similar changes in other subunits of the same molecule, is called cooperativity; Molecules exhibiting cooperativity are also called allosteric molecules; http://upload.wikimedia.org/wikipedia/commons/0/07/Hb-animation2.gif

12. CHMI 2227 - E.R. Gauthier, Ph.D. 12 Quaternary structure Hb is an allosteric protein This phenomenon explains very well the behaviour of Hb in the presence of O 2 :  At low pO 2, all of the Hb subunits in the molecule are in the deoxy form with low affinity for O 2 : they bind O 2 very poorly;  At higher pO 2, one of the 4 subunits binds O 2, changes its conformation to the one with high affinity, and transmits this change in 3D structure to the other 3 subunits;  The other 3 subunits, now having high affinity for O 2, readily bind the molecule and rapidly become saturated. O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 O2O2 Low affinity High affinity Saturated O2O2 O2O2 O2O2

13. CHMI 2227 - E.R. Gauthier, Ph.D. 13 Quaternary structure Why Hb is allosteric, while Myoglobin is not? If Hb behaved like Myoglobin, then most of the Hb molecules would remained tightly bound to O 2 and would not unload O 2 in tissues; Conversely, if myoglobin behaved like Hb, it would readily let go of its O 2, drastically limiting our muscles ability to perform aerobic work;

14. CHMI 2227 - E.R. Gauthier, Ph.D. 14 The Bohr effect The Bohr effect concerns the observed decrease in O 2 binding by hemoglobin when the pH is lowered; This effect explains why hemoglobin binds O 2 in the lungs, and releases it in the tissues; http://www.aw-bc.com/mathews/ch07/fi7p16.htm

15. CHMI 2227 - E.R. Gauthier, Ph.D. 15 Gas exchange In the tissues Tissues Erythrocytes Glucose + O 2 ATP CO 2 H2OH2O Carbonic anhydrase H 2 CO 3 HCO 3 - H+H+ Plasma HCO 3 - (to lungs) Hb-4O 2 Hb-H + 4O 2 H 2 O + Cl -

16. CHMI 2227 - E.R. Gauthier, Ph.D. 16 Gas exchange In the lungs Lungs Erythrocytes CO 2 H2OH2O Carbonic anhydrase H 2 CO 3 HCO 3 - H+H+ Plasma HCO 3 - Hb-4O 2 Hb-H + 4O 2 O2O2 H 2 O + Cl - CO 2 Air CO 2 O2O2