Purification and Characterization of an Esterase Involved in Cellulose Acetate Degradation by Neisseria sicca SB

Abstract

  An esterase catalyzing the hydrolysis of acetyl ester moieties in cellulose acetate was purified 1,110-fold to electrophoretic homogeneity from the culture supernatant of Neisseria sicca SB, which can assimilate cellulose acetate as the sole carbon and energy source. The purified enzyme was a monomeric protein with a molecular mass of 40 kDa and the isoelectric point was 5.3. The pH and temperature optima of the enzyme were 8.0-8.5 and 45°C. The enzyme catalyzed the hydrolysis of acetyl saccharides, p-nitrophenyl esters of short-chain fatty acids, and was slightly active toward aliphatic and aromatic esters. The K_m and V_max for cellulose acetate (degree of substitution, 0.88) and p-nitrophenyl acetate were 0.0162% (716 μM as acetyl content in the polymer) and 36.0 μM, and 66.8 and 39.1 μmol/min/mg, respectively. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate, which indicated that the enzyme was a serine esterase.