Enzyme kinetics Flashcards
(31 cards)
3 characteristics of enzymes in a reaction
- Regenerated during course of reaction 2. Do NOT change difference in free energy between reactants and products (_G: Energy of reactants - energy of products) 3. Do NOT change equilibrium of reactants and products
4 catalytic mechanisms
- Transition state stabilization 2. Bond strain 3. Proximity and orientation 4. Covalent catalysis
Transition state stabilization
Stability prevents transition state from going back to substrate and increases concentration of intermediate and rate of product formation
Catalysis by bond strain
Binding of substrate to enzyme produces bond strain which makes it easier to get to the transition state
Covalent catalysis
Covalent intermediate forms between enzyme and substrate due to orientation of active sites on enzymes
V0
Initial velocity. V0=_[P]/_t Can only be measured at the very beginning of a reaction when very little product made (<5% [S] converted to [P])
Effect of heat and pH on enzymes
Optimal temp and pH ranges where enzymes have the most activity. Outside this range can denature and die
Enzyme saturation
Occurs at high [S], hyperbolic curve
Steady state
At beginning of reaction [ES] builds up but over time reaches state where [ES] remains constant which will persist until almost all of substrate consumed
M-M rate equation 3 assumptions
- [S]»>[E] so only small amount of S bound to E 2. [ES] is unchanged, stays at steady state 3. Initial velocity (V0) used (so reverse reaction not a factor)
M-M equation
v=Vmax_[S]/Km+[S]
Km
[S] at _Vmax Constant for a given enzyme. Estimate of the equilibrium constant for S binding to E. Small Km means tight binding and large Km means weak binding
Vmax
Theoretical maximum velocity. Constant for a given enzyme. To reach Vmax requires ALL of enzyme molecules to have tightly bound substrate
Kcat
Kcat=Vmax/Et (total enzyme) Turnover number. Measure of catalytic activity under saturating substrate conditions. Maximum number of substrate molecules converted to product per enzyme molecule per unit of time. Values range from <1/sec to millions/sec
Lineweaver and Burk plot
Used M-M equation to produce a linear plot of catalyzed reactions. Useful for analyzing enzyme inhibition.
Competitive enzyme inhibitor
Binds the catalytic site and competes with substrate for binding with dynamic equilibrium-like process. Reversible by substrate.
Vmax and Km with competitive inhibitors
Vmax is unchanged (could add a ton of substrate and still get close to Vmax). Km is increased.
Vmax and Km with non-competitive inhibitors
Vmax is decreased. Km is unchanged.
Non-competitive enzyme inhibitor
Binds E or ES complex at site other than the catalytic site. Substrate binding unaltered but ESI cannot form products. Not reversible by substrate.
ACE inhibitors
Angiotensin-converting enzyme inhibitors. Primarily treat HTN and congestive heart failure. Inhibit ACE (component of the blood pressure regulating renin-angiotension system.
What causes high blood pressure?
Overactivation of the renin-angiotension-aldosterone system
Methotrexate
Anti-metabolite and anti-folate drug. Used in cancer treatment, autoimmune diseases by competitively inhibiting the synthesis of DNA, RNA, thymidylates, and proteins.
Aspirin
Supresses production of prostaglandins and thromboxanes due to irreversible inactivation of cyclooxygenase (PTGS) enzyme required for prostaglandin and thromboxane synthesis.
Allosteric enzymes
Most are ogliomeric enzymes (consist of multiple subunits). Located near branch points in metabolic pathways, directing substrates along metobolic paths.
