bioenergetics study of energy transformations in living organisms thermodynamics –1st law:...
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Bioenergetics• Study of energy transformations in living organisms
• Thermodynamics– 1st Law: Conservation of E
• Neither created nor destroyed• Can be transduced into different forms
– 2nd Law: Events proceed from higher to lower E states• Entropy (disorder) always increases
– Universe = system + surrounds
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Bioenergetics(E content of system) H = (useful free E) G + (E lost to disorder) TS
• Gibbs Free Energy: G = H - TS– If G = negative, then rxn is exergonic, spontaneous– If G = positive, then rxn is endergonic, not spontaneous
– Standard conditions (ΔG°’): 25oC, 1M each component, pH 7, H2O at 55.6M
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BioenergeticsA + B <--> C + D
• Rate of reaction is directly proportional to concentration of reactants
• At equilibrium, forward reaction = backward reaction
k1[A][B] = k2[C][D]
• Rearrange:
k1/k2 = ([C][D])/([A][B]) = Keq
• Relationship between ΔG°’ and K’eq is:
G°’ = -2.303 * R * T * log K’eq
If Keq >1, G°’ is negative, rxn will go forwardIf Keq <1, G°’ is positive, rxn will go backward
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• ΔG°’ is a fixed value at standard conditions• ΔG under actual cellular conditions can be different– e.g., for ATP hydrolysis inside a cell, can approach ΔG = -12 kcal/mol
• We will work with ΔG°’ values
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Glutamic acid (Glu) + NH3 --> Glutamine (Gln)G°’=+3.4 kcal/mol
ATP --> ADP + Pi G°’=-7.3 kcal/mol---------------------------------------------------------------------------------------- Glu + ATP + NH3 --> Gln + ADP + Pi
G°’=-3.9 kcal/mol
Glutamyl phosphate is the common intermediate
Coupling endergonic and exergonic rxns
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ATP --> ADP + Pi ΔG°’= -7.3 kcal/molADP + Pi --> ATP ΔG°’= +7.3 kcal/molC(diamond) + O2 --> CO2 ΔG°’= -94.8 kcal/molPEP --> pyruvate + Pi ΔG°’= -14.8 kcal/molC(graphite) + O2 --> CO2 ΔG°’= -94.1 kcal/molP-creatine --> creatine + Pi ΔG°’= -11.0 kcal/molG6-P --> glucose + Pi ΔG°’= -3.0 kcal/mol1,3-BPG --> 3PG + Pi ΔG°’= -12.5 kcal/mol----------------------------------------------------------------------------------------What is ΔG°’ of: PEP + ADP --> pyruvate + ATP
ΔG°’= -7.5----------------------------------------------------------------------------------------
What is ΔG°’ of: G6-P + ADP --> glucose + ATPWhat is ΔG°’ of: P-creatine + ADP --> creatine + ATPWhat is ΔG°’ of: C(s, diamond) --> C(s, graphite)
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Equilibrium vs steady state
• Cells are open systems, not closed systems– O2 enters, CO2 leaves– Allows maintenance of reactions at conditions far from equilibrium
O2
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1) Req’d in small amounts2) Not altered/consumed in rxn3) No effect on thermodynamics of rxn
a) Do not supply Eb) Do not determine [product]/[reactant]
ratio (Keq)c) Do accelerate rate of reaction (kinetics)
4) Highly specific for substrate/reactant5) Very few side reactions (i.e. very “clean”)6) Subject to regulation
No relationship between G and rate of a reaction (kinetics)
Why might a favorable rxn *not* occur rapidly?
Biological Catalysts
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Overcoming the activation energy barrier (EA)• Bunsen burner: CH4 + 2O2 --> CO2 + 2H2O
– The spark adds enough E to exceed EA (not a catalyst)
• Metabolism ‘burning’ glucose– Enzyme lowers EA so that ambient fluctuations in E are sufficient
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Overcoming the activation energy barrier (EA)• Bunsen burner: CH4 + 2O2 --> CO2 + 2H2O
– The spark adds enough E to exceed EA
• Metabolism ‘burning’ glucose– Enzyme lowers EA so that ambient fluctuations in E are sufficient
Catalyst shifts EA line to left <---
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How to lower EA
• The curve peak is the transition state (TS)• Enzymes bind more tightly to TS than to either reactants or products
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How to lower EA
• Mechanism: form an Enzyme-Substrate (ES) complex at active site
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How to lower EA
• Mechanism: form an Enzyme-Substrate (ES) complex at active site– Orient substrates properly for reaction to occur
• Increase local concentration• Decrease potential for unwanted side reactions
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How to lower EA
• Mechanism: form an Enzyme-Substrate (ES) complex at active site– Enhance substrate reactivity
• Enhance polarity of bonds via interaction with amino acid functional groups
• Possibly form covalent bonded intermediates with amino acid side chains
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How to lower EA• Possibly form covalent bonded intermediates with amino acid side chains– Serine protease mechanism:
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How to lower EA• Possibly form covalent bonded intermediates with amino acid side chains– Serine protease mechanism:
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How to lower EA
• Mechanism: form an Enzyme-Substrate (ES) complex at active site– Induce bond strain
• Alter bonding angles within substrate upon binding• Alter positions of atoms in enzyme too: Induced fit
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Induced fit
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Induced fit
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S <--> PAt low [S], rate/velocity is slow, idle time on the enzymeAt very high [S], rate/velocity is maximum (Vmax), enzyme is saturated
V = Vmax [S]/([S] + Km) Km = [S] at Vmax/2
A low Km indicates high enzyme affinity for S(0.1mM is typical)
Enzyme kinetics: The Michaelis-Menten Equation
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Enzyme kinetics: pH and temperature dependence
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Enzyme inhibitors• Irreversible
– Form a covalent bond to an amino acid side chain of the enzyme active site• Block further participation in catalysis
– Example: penicillin• Inhibits Transpeptidase enzyme required for bacterial cell wall synthesis– Weak cell wall = cell burst open
penicillin
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Enzyme inhibitors• Reversible
– Competitive• bind at active site• Steric block to substrate binding
– Km increased– Vmax not affected (increase [S] can overcome)
• Example: ritonavir– Inhibits HIV protease ability to process virus proteins to mature forms
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Enzyme inhibitors• Reversible
– Noncompetitive• Do not bind at active site
• Bind a distinct site and alter enzyme structure reducing catalysis– Km not affected– Vmax decreased, (increase [S] cannot overcome)
NoncompetitiveCompetitive
• Example: anandamide (endogenous cannabinoid)– Inhibits 5-HT3 serotonin receptors that normally
increase anxiety
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Drug discovery• Average cost to market ~ $1B• Average time to market ~13 years• Size of market ~ $289B per year in US (2006)
• S. aureus infections are a problem in hospital settings– Drug targets
• Metabolic rxns specific to bacteria– Sulfa drugs (folic acid biosynthesis)
• Cell wall synthesis– Penicillin, methicillin, vancomycin
• DNA replication, transcription, translation– Ciprofloxacin (DNA gyrase)– Tetracyclins (ribosome)– Zyvox (ribosome)
» Introduced in 2000, resistance observed within 1 year of use