What is enzyme activity and how is it measured?
Enzyme activity is defined as the amount of enzyme that converts 1 μmol of substrate per minute under specified conditions. One unit (U) equals 1 μmol substrate converted per minute. It's measured using spectrophotometric assays or by quantifying product formation over time. See also our Isoelectric Point (pI) Calculator.
How do I convert ΔA/min to enzyme units?
Use the Beer-Lambert law: first convert ΔA/min to concentration change (mM/min) by dividing by extinction coefficient and path length. Then multiply by reaction volume and dilution factor to get μmol/min (enzyme units).
What is specific activity and why is it important?
Specific activity is enzyme activity per unit mass of protein (U/mg). It indicates enzyme purity and efficiency. Higher specific activity means more active enzyme per milligram of protein, which is crucial for enzyme purification and quality assessment.
What dilution factor should I use?
The dilution factor accounts for sample dilution before the assay. If you diluted your enzyme sample 1:10, use dilution factor = 10. If no dilution was made, use 1. This ensures accurate calculation of the original enzyme concentration.
How do I choose between spectrophotometric and product formation methods?
Use spectrophotometric method when you can measure absorbance changes continuously (ΔA/min). Use product formation method when you measure the total amount of product formed over a specific time period. Both give equivalent results when properly calibrated.
What is a typical extinction coefficient value?
Extinction coefficients vary by chromophore: NADH at 340nm = 6,220 M⁻¹·cm⁻¹, p-nitrophenol at 405nm = 18,500 M⁻¹·cm⁻¹. Always use the coefficient specific to your substrate/product and wavelength for accurate calculations.