Abstract
The active site of an N-succinyl-L-trialanine p-nitroanilide-hydrolyzing protease purified from pronase E was investigated. When the enzyme was treated with [3H] diisopropyl phosphofluoridate, an active serine residue of the enzyme was modified, resulting in almost complete loss of the enzymatic activity. The amino acid sequence around the active serine residue (*) was Gly-Asp-Ser*-Gly-Gly. When a histidine residue in the enzyme molecule was destroyed by photooxidation in the presence of methylene blue, its activity decreased up to 15% of the original level. In this treatment, no change was observed either in tyrosine and methionine contents or in ultraviolet absorption spectra. The enzyme was also inactivated by glycine methyl ester in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide but not by p-bromophenacyl bromide. These findings suggest the possibility that serine, histidine, and aspartic or glutamic acid may be involved in the catalytic site of the enzyme to form the charge relay system.
