1. Myoglobin & Hemoglobin
Structure, Function & malfunction of Biomolecules

2. Primary Structure
Sequence of amino acids in a protein connected via peptide linkage.
Example –the enzyme lysozyme:
Lys-Val-Phe-Gly-Arg...Gly-Cys-Arg-Leu
Note: By convention, amino acid sequences are written starting with the amino terminus.

3. Secondary Structure
Regular patterns of relatively small segments of a protein held together mainly by H-bonds
Examples:
-structure
α-helix

4. Tertiary Structure
Overall 3-D shape of a protein. Two basic types are globular and fibrous.
Examples:
Fibrous (Collagen)
Globular (Pepsin)

5. Quaternary Structure Overall 3-D shape of a multi-subunit protein
Example:
Rabbit muscle glycogen phosphorylase

6. Protein Function in Cell
Enzymes
Catalyze biological reactions
Structural role
Cell wall
Cell membrane
Cytoplasm

7. Protein: The Machinery of Life
NH2-Val-His-Leu-Thr-Pro-Glu-Glu-
Lys-Ser-Ala-Val-Thr-Ala-Leu-Trp-
Gly-Lys-Val-Asn-Val-Asp-Glu-Val-
Gly-Gly-Glu-…..

8. Oxygen Transport Proteins
Myoglobin
Exhibits Michaelis-Menten properties
Hemoglobin
Exhibits allosteric properties

9. Myoglobin Single polypeptide with 154(human) amino acids
C774H1224N210O222S5 17,183.8 daltons(human)
8 a helices (A-H)
Located in skeletal & cardiac muscle
[high] in diving mammals like whale & seals

10. pO2 (partial pressure of O2) (Torr)
O2 Binding Curve
Myoglobin has high affinity for O2.
P50 = 2.8 Torr
Allows myoglobin to act as O2 storage reserve.
Releases O2 when pO2 becomes low indicating O2 deprivation.
tissues
arterial pressure
pO2 (partial pressure of O2) (Torr) 20
100
saturation with O2 50
2.8

11. Heme Prosthetic Group Heme (Fe2+) has affinity for O2.
Hematin (Fe3+) cannot bind O2.
Located in crevice where it is protected from oxidation.

13. Oxygen Binding to Myoglobin
distal histidine
O2 binds to only available coordination site on iron atom.
His 93 (proximal his) binds directly to iron.
His 64 (distal his) stabilizes the O2 binding site.
proximal histidine

14. Myoblobin:CO complexes
CO binds tightly; linear.
O2 binds less tightly, bent structure.
Distal His forces bent binding of both, weakens CO binding. Fe C O Fe O

15. Red Blood Cell (Erythrocyte)

16. Model Molecule: Hemoglobin

17. Hemoglobin – Quaternary Structure
a2b2
Two α (141 AA/ α)subunits and two β (146 AA/ β)subunits

18. Heme

19. Hemoglobin Structure
Each polypeptide chain resembles myoglobin tertiary structure but 1˚ sequence varies.
Invariant residues indicate importance of those residues in function.

20. Oxygen Binding Hb exhibits + cooperativity.
Eaton et al. Nature Struct. Biol. 1999, 6, 351

21. O2 Binding to Hemoglobin
Water bound to heme Iron

22. O2 Binding to Hemoglobin
Oxygen bound to heme Iron

23. Hb
T-state deoxy

24. Hb
R-state - oxy

25. Hb Variants HbA2 Embryonic Hb Fetal Hb a2d2 Present in ~2% of adults
Has  affinity for O2
Fetal Hb
a2g2

26. Bohr Effect CO2 pH Some side groups remain protonated at lower pH.
Stabilizes T state and promotes unloading of O2 to active tissues.
Binding of CO2 also stabilizes T state.
CO2 binds to a amino groups.

27. 2, 3-Bisphosphoglycerate
Stabilizes deoxyHb (T state)
Facilitates unloading of O2 in tissue.
100
- BPG
saturation with O2 50
+ BPG 20
100
pO2 (partial pressure of O2) (Torr)

28. 2,3-BPG Binding to Hb

29. High Altitude and BPG
At higher altitudes, the [BPG] increases allowing Hb to unload O2 more easily.

30. Stored Blood & BPG
2,3-BPG becomes depleted in stored blood, so R state of Hb is stabilized.
If BPG depleted blood is used for a transfusion, the R state Hb doesn’t release O2.
Add inosine to stored blood to maintain BPG levels.

31. CO Poisoining CO is “competitive inhibitor” of O2.
Affinity is 200X greater than that of O2.
CO also inhibits unloading O2 of in tissues.

32. Sickle Cell Anemia
Normal red blood cells are round like doughnuts, and they move through small blood tubes in the body to deliver oxygen.
Sickle red blood cells become hard, sticky and shaped like sickles. When these hard and pointed red cells go through the small blood tube, they clog the flow and break apart. This can anemia.

33. The origin of the disease is a small change in the protein hemoglobin
The change in cell structure arises from a change in
the structure of hemoglobin.
A single change in an amino acid causes hemoglobin
to aggregate. a b

34. Scanning electron microscopic image of
Red bllod cells

35. Differences in Red Blood Cells

36. Sickle Cell Hemoglobin
Hemoglobin S
Significant change
in structure caused
by the single mutation

37. L-Glutamic acid (Glu/ E)
L-Valine (Val / V)

38. Sickle Cell Hemoglobin
at 6β
Normal mRNA
GUG CAC CUG ACU CCU GAG GAG AAG
val his leu thr pro glu glu lys
Normal protein
Mutation
(in DNA)
Mutant mRNA
GUG CAC CUG ACU CCU GUG GAG AAG
val his leu thr pro val glu lys
Mutant protein
Glutamate (glu), a negatively charged amino acid, is replaced by valine (val), which has no charge.