Purification and Characterization of a Novel High Molecular Weight Dextransucrase from Acetobacter tropicalis

Abstract

The purification and characterization of extracellular dextransucrase from Acetobacter tropicalis was carried out by using DEAE-Sepharose column chromatography. The enzyme was purified to 16- fold to homogeneity with 8 % recovery. The molecular mass of dextransucrase on SDS-PAGE and non-denaturing SDS-PAGE was found to be 180 and 360 kDa, respectively, suggesting that this enzyme exist in homodimer state in native form. This is a first report on purification of A. tropicalis dextransucrase with such a high molecular weight. Purified dextranuscrase was found to work best with 6 % (w/v) sucrose in acetate buffer (15 mM, pH 5.5) at 37o C temperature. The Km and Vmax values of purified enzyme were found to be 11.5 mM and 5000 U/mg, respectively. The presence of histidine, serine, tyrosine and lysine residues was detected at or near the active sites of enzyme when incubated with amino acid modifiers.