oxygen storage in muscle tissue myoglobin (mb) originally isolated from sperm whales 10x abundance...
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Oxygen Storage in Muscle TissueMyoglobin (Mb)
• Originally isolated from sperm whales• 10X abundance greater in aquatic- than terrestrial-mammals• Mb knockout mice exhibit normal exercise capacity• Detoxification of reactive nitric oxide a signaling molecule
Myoglobin Oxygen Binding Curve
Quantification of Myoglobin’s O2-Binding Behavior
Mb dissociation: MbO2 ↔ Mb + O2
Dissociation constant: K = [Mb][O2]/[MbO2]
Myoglobin-O2 term: [MbO2] = [Mb][O2]/K
Fractional saturation: Y = [MbO2]/([Mb] + [MbO2])
Substitute for MbO2 Y = [Mb][O2]/K/([Mb] + [Mb][O2]/K)
Simplify: Y = [O2]/(K + [O2])
O2 partial pressure: Y = pO2/(K + pO2)
Myoglobin Protein Structure• Single polypeptide chain with 8 alpha helices (A-H)• Prosthetic heme group; only Fe (II)• Central Fe (II) atom bound to 4 porphyrin N atoms • N of His F8 and O2 binds below and above the Fe (II)
Globin EvolutionHomologous proteins with a common ancestor
Genetic mutations with invariant residues
Embryonic hemoglobin
Adult hemoglobin
Hemoglobin: Oxygen Transport in MammalsHemoglobin features:
• Located in red blood cells; 4 subunits• Cooperative oxygen binding (allosteric protein)• Deoxygenated T-form and Oxygenated R-form
Hemoglobin Cooperativity Enhances O2 Delivery
• Proximal His binds Fe (II) • Distal His disfavors CO binding• O2 → Fe (II) shift from out of plane to porphyrin plane
Limitations with Hemoglobin as an oxygen transporter?
Conformational Change in
Hemoglobin
Which is the T and R configuration?
Is this enzyme conversion concerted or sequential?
2,3-Bisphosphoglycerate Stabilizes Hemoglobin
T Configuration
Central cavity size difference T versus R?
Amino acids that comprise the central cavity?
How does 2,3-BPG affect T versus R configuration?
Hemoglobin Central Cavity with 2,3-Bisphosphoglycerate
Bohr Effect: Protons and Carbon Dioxide Promote the T State
• Stabilizing the T state increases hemoglobin O2 release
• T - more sigmodial
• R - more hyperbolic
• T ↔ R + H+
T-State Stabilization by Salt Bridge Formation
How is CO2 linked with H+ concentration?
Carbon Dioxide Acidifies the Hemoglobin Environment
Tissue-Specific Hemoglobin Affinity for Oxygen
What is the favorable hemoglobin state (T/R) in capillaries near muscle tissue and lungs?
How must fetal hemoglobin oxygen binding vary from the mother?
Adjusted Hemoglobin-Oxygen Affinity with Environmental Changes
Fetal hemoglobin (α2γ2) has a Ser substitution for His
Two less positive charges in central cavity means what?
Adjusted Hemoglobin-Oxygen Affinity with Environmental Changes
Hemoglobin Mutation: Sickle-Cell Anemia
Sickle-Cell Features
• Lower hemoglobin (Hb) solubility for Hb S than Hb A (normal)
• Amino acid substitution Glu → Val
• Reduced T-state solubility
• O2 affinity and allosteric properties unaffected
• Heterozygotes asymptomatic
Do changes in the oxygen-binding prosthetic group preclude cooperative O2 binding?
Copper Oxygen Carrier Present in Lobster Hemoglobin
Site-Directed Mutagenesis Experimentation
• Convert proximal His → Gly
• Exogenous imidazole binds with porphyrin iron
Would modified hemoglobin (i)exhibit cooperative binding?
(ii) generate Fe (III)?
Substitution for 2,3-Bisphosphoglycerate in
Bird Hemoglobin
Which compound(s) are well suited to bind in the central cavity of bird hemoglobin?
Physiological [CO2] and [2,3-BPG] at pH = 7
Curve matching:
Increase in [CO2]
Increase in 2,3-BPG
Increase in pH
Loss of 4° structure
Hemoglobin Fractional Saturation Curve Shift
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