PURIFICATION, CHARACTERIZATION, AND SEQUENCE-ANALYSIS OF 2-AMINOMUCONIC 6-SEMIALDEHYDE DEHYDROGENASE FROM PSEUDOMONAS PSEUDOALCALIGENES JS45

2-Aminonumconic B-semialdehyde is an unstable intermediate in the biod egradation of nitrobenzene and 2-aminophenol by Pseudomonas pseudoalca ligenes JS45, Previous work has shown that enzymes in cell extracts co nvert 2-aminophenol to 2-aminomuconate in the presence of NAD(+), In t he present work, 2-aminomuconic semialdehyde dehydrogenase was purifie d and characterized, The purified enzyme migrates as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molec ular mass of 57 kDa, The molecular mass of the native enzyme was estim ated to be 160 kDa by gel filtration chromatography, The optimal pH fo r the enzyme activity was 7.3. The enzyme is able to oxidize several a ldehyde analogs, including 2-hydroxymuconic semialdehyde, hexaldehyde, and benzaldehyde. The gene encoding 2-aminomuconic semialdehyde dehyd rogenase was identified by matching the deduced N-terminal amino acid sequence of the gene with the first 21 amino acids of the purified pro tein. Multiple sequence alignment of various semialdehyde dehydrogenas e protein sequences indicates that 2-aminomuconic 6-semialdehyde dehyd rogenase has a high degree of identity with 2-hydroxymuconic 6-semiald ehyde dehydrogenases.