I understand that enzyme concentration doesn't affect Km since the Michaelis–Menten equation is based on the steady state approximation which requires high levels of substrate compared to enzyme (that's why changing enzyme concentration is negligible compared to substrate concentration and therefore there is no apparent change in Km).

But what about change of temperature or pH? It is known that change in those parameters can change enzyme activity, but does this actually change kinetic parameters like Km?

Why should I think not? Because the same explanation that says that change in enzyme concentration doesn't affect Km should apply here. There is a lot of substrate anyway and therefore there should not be an apparent change in the Km?

Another issue which makes me think so: In all the biochemistry literature that I have encountered Km and Kcat appears for each enzyme with no indication of the temperature or pH for which those values were observed (not in Lehninger, nor Wikipedia or other textbooks).

I have not been able to find a detailed consideration of this point.

  • 1
    $\begingroup$ Wikipedia: “The value of Km is dependent on both the identity of enzyme and that of the substrate, as well as conditions such as temperature and pH.” $\endgroup$
    – acvill
    Jan 2, 2022 at 16:45

1 Answer 1


Michaelis constant is defined as $$K_\textrm{M} = \frac{k_{-1}+k_2}{k_1},$$ where $k$s represent the rate constants for the reactions $$\mathrm{E+S} \;\;\underset{k_{-1}}{\overset{k_1}{\longrightleftharpoons}} \;\;\mathrm{ES}\; \overset{k_2}{\longrightarrow}\;\mathrm{E+P}.$$ Rate constants can generally be expressed by the Arrhenius equation: $$k(T) = A(T)\exp\left(-\frac{E_a}{RT}\right).$$ Because the parameters $A(T)$ (pre-exponential factor) and $E_a$ (activation energy) are generally not equal for all three reactions, the change in temperature does change the value of $K_\rm{M}$.

When measuring the enzyme kinetics and calculating $K_\rm{M}$, one has to explicitly state at which temperature the experiments were performed. This is probably not mentioned afterwards in the textbook because it is not of significant importance to the surrounding text and concepts.

Here you can find an article example with the measurements of enzyme kinetics at different temperatures. You can clearly see the change in $K_\rm{M}$.

Enzyme Kinetics


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