Enzyme Kinetics Overview

Study Notes

Enzyme Kinetics

Enzyme kinetics is the study of the rates of enzyme-catalyzed reactions and the factors affecting those rates.
The word "kinetics" comes from the Greek word "kinetos," meaning "moving."
Biochemistry studies the quantitative measurement of enzyme-catalyzed reactions and the systematic analysis of factors affecting their rates.

Simple Reaction

A simple reaction involves a reactant (substrate, S) transforming into a product (P).
The rate, V, represents the quantity of substrate that disappears or product that appears per unit of time.

Experiment

Enzyme (E) concentration is held constant ([E] is fixed)
Substrate concentration ([S]) is varied (increased)
Measure the initial velocity (Vo)
Initial velocity, Vo, directly relates to substrate concentration [S].
The initial velocity (Vo) directly relates to substrate concentration at low [S].
Initial velocity (Vo) equals maximum velocity (Vmax) at high [S].

Michaelis-Menten Equation

A simple kinetic model proposed in 1913 by Leonor Michaelis and Maud Menten.
The model posits that an enzyme-substrate complex (ES) is a necessary intermediate in enzyme reactions.
E = Enzyme
S = Substrate
ES = Enzyme-substrate complex
P = Product
k1, k-1, k2 = rate constants

Michaelis-Menten Kinetics

The first step involves reversible and quick binding of enzyme (E) and substrate (S) to form a non-covalent enzyme-substrate complex (ES).
The second step involves a chemical transformation of the ES complex that leads to the formation of the enzyme (E) and the product (P).
Velocity (V) is directly proportional to the enzyme-substrate complex (ES) concentration.
Most enzyme reactions follow Michaelis-Menten kinetics.
The enzyme can either be free [E] or bound in the ES complex.
Enzyme concentration ([E]) is negligible compared to substrate concentration ([S]) at sufficiently high [S].
The enzyme works at full capacity (V = Vmax) at high [S].
The concentration of product [P] is near zero at the beginning of the reaction, making k-2 close to zero.

The Michaelis-Menten Equation

Equation is based on a simple reversible reaction (S <-> P) with E+S <=> ES <=> E+P.
The reverse reaction (P->S) is not considered in calculating initial rates.
The enzyme-substrate complex (ES) concentration remains constant due to continuous production and breakdown at the same rate.
V₀ = Vmax[S] / (Km + [S])
V₀ - initial velocity of reaction
Vmax - maximum velocity of reaction
[S] - substrate concentration
Km - Michaelis constant.

Michaelis-Menten Constant (Km)

Km is a constant for any enzyme and substrate pair.
Km is independent of enzyme or substrate concentration.
Km values relate to the enzyme's affinity for the substrate.
Small Km means tighter binding; whereas, a large Km indicates weaker binding.
Km is the substrate concentration at which the reaction velocity is half the maximum reaction velocity (½ Vmax).

Catalytic Rate Constant (kcat)

kcat = Vmax / [ET]; turnover number
Turnover number: number of catalytic cycles that each enzyme active site undergoes per unit time.
kcat/Km = catalytic efficiency; represents the enzyme's overall ability to convert substrate to product.
Upper limit kcat/Km is diffusion controlled (about 10⁸-10⁹ M⁻¹s⁻¹).

Using Enzyme Kinetic Graphs

Direct measurement of Vmax and Km requires high substrate concentrations to achieve saturation.
Lineweaver-Burk plots are linear representations of the Michaelis-Menten equation.

Lineweaver-Burk Plot

Created in 1934. Is a useful graph for analyzing the Michaelis-Menten equation, such as different forms of enzyme inhibition. It's a double reciprocal plot.
Helpful for extrapolating Vmax before powerful computers and nonlinear regression were widely available.

Description

This quiz covers the essential concepts of enzyme kinetics, focusing on the rates of enzyme-catalyzed reactions and how various factors influence these rates. Key topics include simple reactions, experiments measuring initial velocities, and the Michaelis-Menten equation.