Chemical modification by 2,4,6-trinitrobenzenesulfonic acid (TNBS) of an essential amino group in 3-ketovalidoxylamine A C-N lyase.

3-Ketovalidoxylamine A C-N lyase of Flavobacterium saccharophilum is a monomeric protein with a molecular weight of 36000, and contains 32 amino groups and no cysteine or cystine residues. The enzyme was inactivated by 2,4,6-trinitrobenzenesulfonic acid (TNBS) following pseudo-first order kinetics. Substrate of the lyase, p-nitrophenyl-3-ketovalidamine, protected the enzyme against the inactivation, suggesting that the modification occurred at or near the active site. Although several amino groups were modified by TNBS, a plot of log (reciprocal of the half-time of inactivation) versus log (concentration of TNBS) suggested that one amino group has an essential role in catalysis.