Transcript Essential Biochemistry

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
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 Evolution
Homologous proteins with a
common ancestor
Genetic mutations with
invariant residues
Adult
hemoglobin
Embryonic
hemoglobin
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
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?
Hemoglobin Central Cavity with
2,3-Bisphosphoglycerate
How does 2,3-BPG affect T versus R configuration?
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?
Adjusted Hemoglobin-Oxygen Affinity
with Environmental Changes
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?
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
Copper Oxygen
Carrier Present in
Lobster Hemoglobin
Do changes in the
oxygen-binding
prosthetic group preclude
cooperative O2 binding?
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 2,3Bisphosphoglycerate in
Bird Hemoglobin
Which compound(s)
are well suited to bind
in the central cavity of
bird hemoglobin?
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