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Myoglobin & Hemoglobin
Structure, Function & malfunction of Biomolecules
Primary Structure
Sequence of amino acids in a protein connected via peptide linkage.
Example –the enzyme lysozyme:
1 2 3 4 5 126 127 128 129
Lys-Val-Phe-Gly-Arg...Gly-Cys-Arg-Leu
Note: By convention, amino acid sequences are written starting with the amino terminus.
Secondary Structure
Regular patterns of relatively small segments of a protein held together mainly by H-bonds
α-helix -structure
http://www.ultranet.com/~jkimball/BiologyPages/S/SecondaryStructure.html
Examples:
Tertiary Structure
Overall 3-D shape of a protein. Two basic types are globular and fibrous.
Globular (Pepsin) Fibrous (Collagen)
Examples:
Quaternary Structure
Overall 3-D shape of a multi-subunit protein
Rabbit muscle glycogen phosphorylase
http://bmbiris.bmb.uga.edu/wampler/tutorial/prot4.html
Example:
Protein Function in Cell
1. Enzymes Catalyze biological reactions
2. Structural role Cell wall Cell membrane Cytoplasm
Protein: The Machinery of LifeNH2-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-…..
Oxygen Transport Proteins Myoglobin
Exhibits Michaelis-Menten properties
Hemoglobin Exhibits allosteric properties
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
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.
tissu
es
arte
rial p
ress
ure
pO2 (partial pressure of O2) (Torr)
20 100
satu
ratio
n w
ith O
2
100
50
2.8
N
N N
N
HO O
Fe
Heme (Fe2+) has affinity for O2.
Hematin (Fe3+) cannot bind O2.
Located in crevice where it is protected from oxidation.
Heme Prosthetic Group
Oxygen Binding to Myoglobin
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.
http://cwx.prenhall.com/horton/medialib/media_portfolio/text_images/FG04_44.JPG
distal histidine
proximal histidine
• CO binds tightly; linear.• O2 binds less tightly, bent structure.
• Distal His forces bent binding of both, weakens CO binding.
Fe
OO
Fe
CO
Myoblobin:CO complexes
Red Blood Cell (Erythrocyte)
Model Molecule: Hemoglobin
Hemoglobin – Quaternary Structure
Two α (141 AA/ α)subunits and two β (146 AA/ β)subunits
a2b2
Heme
Hemoglobin Structure
Each polypeptide chain resembles myoglobin tertiary structure but 1˚ sequence varies.
Invariant residues indicate importance of those residues in function.
Oxygen Binding Hb exhibits + cooperativity.
Eaton et al. Nature Struct. Biol. 1999, 6, 351
Water bound to heme Iron
O2 Binding to Hemoglobin
Oxygen bound to heme Iron
O2 Binding to Hemoglobin
HbT-state deoxy
Hb R-state - oxy
Hb Variants
http://oregonstate.edu/instruction/bb450/stryer/ch10/Slide27.jpg
HbA2
a2d2 Present in ~2% of adults
Embryonic Hb a2e2
Has affinity for O2
Fetal Hb a2g2
Has affinity for O2
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. http://cwx.prenhall.com/horton/medialib/media_portfolio/text_images/FG04_50.JPG
2, 3-Bisphosphoglycerate Stabilizes deoxyHb (T
state)
Facilitates unloading of O2 in tissue.
O O
O
O-
P
O
O-
O-
P
O
O-
-O
2, 3-bisphosphoglycerate
pO2 (partial pressure of O2) (Torr)
20 100
satu
ratio
n w
ith O
2
100
50
- BPG
+ BPG
2,3-BPG Binding to Hb
http://oregonstate.edu/instruction/bb450/stryer/ch10/Slide26.jpg
High Altitude and BPG
At higher altitudes, the [BPG] increases allowing Hb to unload O2 more easily.
http://www.bio.davidson.edu/Courses/anphys/1999/Yusi/dpgoxyhbgraph.jpg
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.
CO Poisoining
CO is “competitive inhibitor” of O2. Affinity is 200X greater than that of O2.
CO also inhibits unloading O2 of in tissues.
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.
The origin of the disease is a small change in the protein hemoglobin
The change in cell structure arises from a change inthe structure of hemoglobin.
A single change in an amino acid causes hemoglobinto aggregate.
ab
Scanning electron microscopic image of Red bllod cells
Differences in Red Blood Cells
Sickle Cell Hemoglobin
Significant change
in structure caused
by the single mutation
Hemoglobin S
L-Valine(Val / V)
L-Glutamic acid(Glu/ E)
Sickle Cell Hemoglobin
GUG CAC CUG ACU CCU GAG GAG AAGval his leu thr pro glu glu lys 1 2 3 4 5 6 7 8
GUG CAC CUG ACU CCU GUG GAG AAGval his leu thr pro val glu lys 1 2 3 4 5 6 7 8
Mutation (in DNA)
Normal mRNA
Normal protein
Mutant mRNA
Mutant protein
Glutamate (glu), a negatively charged amino acid, is replaced by valine (val), which has no charge.
at 6β