GraphPad Curve Fitting Guide

Key concepts: Substrate vs. velocity

Key concepts: Substrate vs. velocity

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Key concepts: Substrate vs. velocity

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Fitting a substrate-velocity curve

The simplest experiment in enzyme kinetics is to vary the substrate concentration and measure enzyme velocity (also called enzyme activity).

The standard way to fit these data is to fit the Michaelis-Menten model to determine the Vmax (maximum enzyme velocity) and its Km (the concentration of substrate needed to get half-maximal velocity.

The Vmax equals the product of the concentration of active enzyme sites times the turnover rate, kcat. This is the number of substrate molecules each enzyme site can convert to product per unit time. If you know the concentration of enzyme, you can fit the curve to determine kcat and Km. The curve will be identical to the Michaelis-Menten fit.

If the enzyme has cooperative subunits, the graph of enzyme velocity as a function of substrate concentration will appear sigmoidal. Prism offers one empirical equation for fitting sigmoidal substrate-velocity curves.