Quiz 12 Flashcards
kinetics
- the branch of science concerned with the rates of reactions
enzyme kinetics
seeks to determine the initial and maximal reaction velocity that enzymes can attain and the binding affinities for substrates and inhibitors
simple rate equations describe the progress of
first and second order reactions
michaelis-menten equation
relates the initial velocity of a reaction to the maximal reaction velocity and the Michaelis constant for a particular enzyme and substrate
enzyme catalytic efficiency is expressed as
Kcat/Km
bisubstrate reactions can occur by
ordered or random sequential mechanisms or by pingpong mechanism
line weaver- Burk plot
can be used to present kinetic data and to calculate values for Km and Vmax. useful for analysis of inhibitors
rate law
mathematical relationship between the reaction rate, or velocity, “n” and concentration of reactants
v = Vmax[s]/Km+[s]
rate or velocity
the amount of A consumed or P formed per unit time [t]
rate constant k
proportionality constant in the rate equation
simple first order rxns display
a linear plot of the substrate [a] and product [p]
the order for any reactant is given by its
exponent in the rate equation
enzyme catalyzed rxns are more complex than
chemically catalyzed rxns
Vo [initial velocity] is a number equal to
the slope at the beginning of each time course
enzyme catalyzed rxns
- at low [s] the rate is proportional to s as in first-order rxn : n=k [S]
- at high [s], the enzyme reaction approaches zero order kinetics : n= Vmax
rate of formation of ES: k1
rate of disassociation: k-1
rate of product formation : k2
initial velocity is plotted as
a function of substrate conc
Leonor Michaelis and Maud mentens theory
- assumes formation of enzyme-substrate complex [ES]
- assumes the [ES] is constant, “steady-state assumption”
- assumes that at high [s] the breakdown of ES [k2] to form product P, is the rate limiting step, equal to the catalytic rate constant , Kcat
steady-state assumption
- no change in the conc of the ES complex
- rapid flux means the ES complex either dissociates or results in the product formation, as rapidly as new ES complexes are forming
lineweaver-burk plots
offer another useful form rate of law
- begin with n= Vmax [s]/ (Km +[s]) and take reciprocal of both sides
- distinguish between types of bisubstrate reactions
Km
- substrate conc at 1/2 Vmax
2. K-1+K2/ K1
small km means little dissociation [tight binding]
large km means lots of dissociation [little binding]
Vmax
- constant for each enzyme equal to Kcat [E]t
- the theoretical maximal rare of the reaction [never achieved in reality ]
turnover number
Kcat is the turnover number
K2=Kcat= Vmax/ET
catalytic efficiency , Kcat/Km
Kcat/Km
- a way to score how well an enzyme catalyzes a reaction
- upper limit is the diffusion limit approaching 1x10^9, the rate at which E and S encounter each other by diffusion
enzyme substrate recognition and catalysis are greatly dependent on
pH
rates of enzyme-catalyzed reactions generally increase with
increasing temp.
- at temps above 50 to 60c enzymes typically show a decline in activity
2 effects:
1. enzyme rate typically doubles in rate for every 10C as long as the enzyme is stable and active
2. at higher temps, the protein becomes unstable, and denaturation occurs
bisubstrate reactions may be sequential or single displacement reactions or double displacement [pingpong] reactions
- single displacement aka sequential can be two distinct classes
a. random
b. ordered - double displacement [pingpong] rwactions
involves a covalent intermediate
double displacement [ping-pong] reactions proceed via formation of a
covalently modified enzyme intermediate, E’ product [p] of the reaction with A is released prior to the rxn of the enzyme with the second substrate
- characterized by parallel lines
reversible inhibitors may
bind at the active site or at some other site
enzymes may also be inhibited in an
irreversible manner
- penicillin is an irreversible, suicide inhibitor
competitive
raises Km with no change in Vmax
noncompetitive
decreases Vmax with no change in Km
mixed noncompetitive
alters both Km and Vmax
uncompetitive
alters both Km and Vmax but with same slope, Km/Vmax
assessment of kinetic mparamenetes using the michaelis-menten equation and line weaver-burk plots allows for
- characterization of enzymes, identification of bi-substrate reactions and characterization of inhibitors
kinetics
- kinetics analysis is a deductive reasoning tool
- kinetics cannot prove a reaction mechanism
- kinetics can only rule out various alternative hypotheses because they dont fit the data
