Abstract

An intercellular carboxylesterase (EC 3.1.1.1) was isolated from Arthrobacter viscosus NRRL B-1973 and purified to homogeneity. Purification of extracted enzyme was achieved by ammonium sulfate precipitation, followed by separation with Phenyl-Sepharose, DEAE-Sepharose, and Sepharose CL-6B. This carboxylesterase exhibits a specific activity of 55.4*10^-6 moles min^-1 based on the hydrolysis of p-nitrophenol acetate at pH 7.4 and 30° C. The apparent molecular mass was 16.7±0.4 kDa as determined by SDS-PAGE. The isoelectric point was estimated to be 5.6 and optimum activity for the enzyme was found at pH 7.4 and 40° C. The enzyme deacetylated xanthan, alginate, glucose pentaacetate, cellobiose octaacetate and the native exopolysaccharide produced by A. viscosus. The enzyme also deacetylated a variety of low molecular weight esters. In addition, the enzyme can also catalyze transesterification from selected acyl donors to polymeric acceptors including cellulose. One and two dimensional ¹³C / ¹H NMR studies on cellobiose indicate acetylation at the O(2) and O(3) but not O(6) positions. CP/MAS NMR was used to demonstrate the in vitro acetylation by this enzyme of microcrystalline cellulose dispersed in water.