Chromogenic Substrate (Molecular Biology)

A chromophore is any light-absorbing group (see Absorption Spectroscopy), and a chromogenic substrate is one that is acted on by an enzyme so as to increase or decrease the absorption of light at a particular wavelength as substrate is converted to product. An example of a naturally occurring chromogenic substrate is NADH, which absorbs light strongly at 340 nm. The absorbance decreases as NADH is oxidized by a pyridine nucleotide-dependent dehydrogenase, because NAD does not absorb substantially at 340 nm. Artificial chromogenic substrates have been used extensively for kinetic studies on a range of enzymes. p-Nitrophenylphosphate is hydrolyzed by phosphatases to release p-nitrophenol, which has a yellow color in alkaline solution. The increase in absorbance with hydrolysis is measured at 400 nm. p-Nitrophenylesters are used in a similar manner to measure the activity of esterases. Phenazine methosulfate is used as a convenient means of determining the activity of flavoprotein enzymes, as the oxidized form of the electron acceptor is yellow and the reduced form is colorless. An extensive list of artificial, chromogenic substrates has been recorded (1).