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Oxygen Storage in Muscle Tissue

Myoglobin (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. Oxygen Storage in Muscle Tissue.

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Oxygen Storage in Muscle Tissue

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  1. Myoglobin (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 Oxygen Storage in Muscle Tissue

  2. Quantification of Myoglobin’s O2-Binding Behavior Mb dissociation: MbO2 ↔ Mb + O2Dissociation constant: K = [Mb][O2]/[MbO2] Myoglobin-O2 term: [MbO2] = [Mb][O2]/K Fractional saturation: Y = [MbO2]/([Mb] + [MbO2])Substitute for MbO2Y = [Mb][O2]/K/([Mb] + [Mb][O2]/K)Simplify: Y = [O2]/(K + [O2]) O2 partial pressure: Y = pO2/(K + pO2) Myoglobin Oxygen Binding Curve

  3. 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)

  4. Globin Evolution Homologous proteins with a common ancestor Genetic mutations with invariant residues Adult hemoglobin Embryonic hemoglobin

  5. Hemoglobin: Oxygen Transport in Mammals • Hemoglobin features: • Located in red blood cells; 4 subunits • Cooperative oxygen binding (allosteric protein) • Deoxygenated T-form and Oxygenated R-form

  6. 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?

  7. Conformational Change in Hemoglobin Which is the T and R configuration? Is this enzyme conversion concerted or sequential?

  8. 2,3-Bisphosphoglycerate Stabilizes Hemoglobin T Configuration Central cavity size difference T versus R? Amino acids that comprise the central cavity?

  9. Hemoglobin Central Cavity with 2,3-Bisphosphoglycerate How does 2,3-BPG affect T versus R configuration?

  10. 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+

  11. T-State Stabilization by Salt Bridge Formation How is CO2 linked with H+ concentration?

  12. Carbon Dioxide Acidifies the Hemoglobin Environment

  13. Tissue-Specific Hemoglobin Affinity for Oxygen What is the favorable hemoglobin state (T/R) in capillaries near muscle tissue and lungs?

  14. Adjusted Hemoglobin-Oxygen Affinity with Environmental Changes How must fetal hemoglobin oxygen binding vary from the mother?

  15. 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?

  16. 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

  17. Copper Oxygen Carrier Present in Lobster Hemoglobin Do changes in the oxygen-binding prosthetic group preclude cooperative O2 binding?

  18. Site-Directed Mutagenesis Experimentation • Convert proximal His → Gly • Exogenous imidazole binds with porphyrin iron • Would modified hemoglobin • exhibit cooperative binding? • (ii) generate Fe (III)?

  19. Substitution for 2,3-Bisphosphoglycerate in Bird Hemoglobin Which compound(s) are well suited to bind in the central cavity of bird hemoglobin?

  20. Hemoglobin Fractional Saturation Curve Shift Curve matching: Increase in [CO2] Increase in 2,3-BPG Increase in pH Loss of 4° structure Physiological [CO2] and [2,3-BPG] at pH = 7

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