Download ch3a FA11 - Cal State LA

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
Bioenergetics
(E content of system) H = (useful free E) G + (E lost to disorder) TS
• Gibbs Free Energy: G = H - TS
– 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
Bioenergetics
A + 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 forward
If Keq <1, G°’ is positive, rxn will go backward
• 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
Coupling endergonic and exergonic rxns
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
ATP --> ADP + Pi
ΔG°’= -7.3 kcal/mol
ADP + Pi --> ATP
ΔG°’= +7.3 kcal/mol
C(diamond) + O2 --> CO2
ΔG°’= -94.8 kcal/mol
PEP --> pyruvate + Pi
ΔG°’= -14.8 kcal/mol
C(graphite) + O2 --> CO2
ΔG°’= -94.1 kcal/mol
P-creatine --> creatine + Pi
ΔG°’= -11.0 kcal/mol
G6-P --> glucose + Pi
ΔG°’= -3.0 kcal/mol
1,3-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 + ATP
What is ΔG°’ of: P-creatine + ADP --> creatine + ATP
What is ΔG°’ of: C(s, diamond) --> C(s, graphite)
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
Biological Catalysts
1) Req’d in small amounts
2) Not altered/consumed in rxn
3) No effect on thermodynamics of rxn
a) Do not supply E
b) Do not determine [product]/[reactant] ratio (Keq)
c) Do accelerate rate of reaction (kinetics)
4) Highly specific for substrate/reactant
5) 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?
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
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
<---
How to lower EA
• The curve peak is the transition state (TS)
• Enzymes bind more tightly to TS than to either reactants or products
How to lower EA
• Mechanism: form an Enzyme-Substrate (ES) complex at active site
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
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
How to lower EA
• Possibly form covalent bonded intermediates with amino acid side chains
– Serine protease mechanism:
How to lower EA
• Possibly form covalent bonded intermediates with amino acid side chains
– Serine protease mechanism:
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
Induced fit
Induced fit
Enzyme kinetics: The Michaelis-Menten Equation
S <--> P
At low [S], rate/velocity is slow, idle time on the enzyme
At 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: pH and temperature dependence
Enzyme inhibitors
• Irreversible
– Form a covalent bond to an amino acid side
chain of the enzyme active site
• Block further participation in catalysis
penicillin
– Example: penicillin
• Inhibits Transpeptidase enzyme required
for bacterial cell wall synthesis
– Weak cell wall = cell burst open
Enzyme inhibitors
• Reversible
• Example: ritonavir
– Competitive
– Inhibits HIV protease ability to
• bind at active site
process virus proteins to mature
• Steric block to substrate binding
forms
– Km increased
– Vmax not affected (increase [S] can overcome)
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)
Competitive
• Example: anandamide
(endogenous cannabinoid)
– Inhibits 5-HT3 serotonin
receptors that normally
increase anxiety
Noncompetitive
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