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28 Cards in this Set
- Front
- Back
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Biological role of Mb
Biological role of Hb |
Mb: Stores oxygen in muscle
Hb: transport protein (oxygen from lungs to tissues), nearly 1/7th of protein is RBC |
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Similarity of Mb and Hb
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Same prosthetic group (heme)
Same ligand (oxygen) 2nd and 3rd structure very similar (primary sequences have some identical and similar sequences) |
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How many domains are in hemoglobin? How many in myoglobin?
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Hemoglobin: 4 domains (tetramer)
Myoglobin:1 domain (monomer) |
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Type of subunits for myoglobin/hemoglobin
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Alpha helices for myoglobin, alpha2x2 forHb (X=isoforms for beta, gamma, delta
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Main difference between myoglobin and hemoglobin folds
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Little alpha helix
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X isoforms for Hb tetramer
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alpha2beta2 human adult
alpha2gamma2 human fetal alpha2delta2 human adult minor form |
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Can Mb tetramize with Hb? What is in a hemoglobin tetramer?
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No Mb can't, beta4 assembles but does not function, mix of Hbalpha and Hbbeta=alpha2beta2
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Hemoglobin ligands
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First-heme group,bound tightly, prosthetic group
Second-dioxygen molecule, more frequent association/disassociation, oxygen bind to heme group |
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Why does protein bind to heme?
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1. Free heme more affinity to CO than O2. Protein hinders CO binding
2.Inhibits reaction of iron oxidation of iron in the heme (competitive binding) |
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How to detect oxygen binding in hemoglobin/myoglobin
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Blue:deoxy
Red:oxy Measure absorbance Hb + O2 -HbO2 Kd=dissociation Simple hyperbolic binding curve Y=[O2]/(Kd + [O2]) (sigmoidal on log scale) HEMOGLOBIN USUALLY USED IN LINEAR SCALE |
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What time of binding for Hb (based off observed binding curve)
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Positive cooperative binding (n=3, when it should be 4, not exactly stoichiometry)
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Which protein binds O2 more tightly?
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Myoglobin: in muscle-need to grab oxygen from hemoglobin
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Which is more hyperbolic? sigmoidal? what is the x label?
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Myoglobin:hyperbolic
Hemoglobin: sigmoidal X-gaseous pressure of oxygen PO2 |
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What is Hb's positive cooperativity due to?
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Conformation changes: first 2 O2 bind, causes conformational change shift, last 2 O2 bind with higher affinity
Reverse process in muscle |
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Is Hb's conformational change really 2state salt bridge?
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Too simplistic: two states are end points of continuum
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Hb allostery: BGP. Tell me about it.
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2,3-bisphosphoglycerate (synthesized in erythrocytes from glucose)
1BPG per Hb tetramer, between 2 beta chains,nearly every Hb bound Negatively charged ligand, surrounded by positive charged sidechains Binds preferentially to de-oxy form (does not bind to oxy form) Lost during blood storage (BPG disappears if it is on shelf too long--unregulated in hypoxia, deficiency in oxygen delivery to tissues) |
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Does Hb allostery increase or decrease O2affinity? What happens to Hb conformational equilbrium
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O2 affinity reduced
Without BPG, curve looks more like Mb Pulls to the left (from oxyHb) BPG facilitates the conformational change to deoxy Hb |
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Since Hb allostery weakens O2 affinity, how does it affect cooperativity? What do the curves look like?
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Greatly enhances cooperativity. Without BPG,curve would look more like Mb (not enough deoxy to have significant cooperativity)
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Is BPG binding weaker or stronger in fetal Hb?
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Weaker--does not bind well (His2Ser substitution) Ser has no charge, losing stabilization because charge is lost (binding affinity diminished)
Oxygen is transferred from mother, weaker BPG binding results in strong O2 binding, Hb gotta work just as hard as Mb to bind to O2 (results in strong O2 binding) |
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Do allosteric factors strengthen or weaken O2 affinity?
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Weaken!
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What does the following do to Hb, and where do they bind: H+, CO2
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H+=weakens O2 binding, specific binding sites unknown
Co2=weakens O2 binding, covalent and reversible binding at N-terminal amines (CO is pathological, CO2 is normal) |
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What is moonlighting?
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Change of NO to NO3-, facilitated by reduced MbO2 binding to NO, with NO reacting with O2 to NO3-
Happens in both Mb and Hb |
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How does altitude changes affect Hb loading?
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Higher altitudes--moves oxygen back in sigmoidal curve (initial percent saturation is lower)
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What happens to Hb curve as pH decreases? (acidonic) Is O2 affinity higher or lower? Is the midpoint higher or lower? Rationalize metabolism
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Protein is an ALLOSTERIC effector, Lowers the affinity to O2, The curve moves more to the right, More oxygen is delivered to tissue, as more oxygen is needed for metabolism
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What happens to Hb curve in the presence of drugs that oxidize Fe2+? Does the midpoint change?
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Curve plateaus at lower rate (Fraction bound can never approach 1. since there is oxygenbounded Hb as a result of the drug
Midpoint doesn't change (same Kd) |
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Genetic diseases of Hb: thalessemias, hemoglobinopathies
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1. required quaternary structure (heterozygous defects vs homozygous defects)
2. thalessemias: loss of one subunit, can't make alpha4beta4, results in low concentration of Hb tetramer (anemia) 3. Point mutations: hemoglobinopathies: i.e. sickle cell anemia |
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Hb Kansas
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Curve shifts to right, looks more like myoglobin (cooperativity decreased, oxygen affinity decreased)
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Synthetic blood: pros and cons
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Pro: purified from blood sources, no antigens, faster recovery from BPG depletion
Con: free Hb cannot withstand stress/pressure of circulation (pressure is a denaturant)--dissociates the tetramer to dimer) Con: Free hb scavenges blood vessels:: moonlighting:: NO is a vasodilator |
