Studies on the Synthesis of New Oligosaccharide by Bacillus subtilis Levansucrase and Biological activity of the Synthesized Sugar.

点击翻译按钮获取中文摘要

Levansucrase was obtained as an electrophoretically pure state from cultured cells of Bacillus subtilis grown on sucrose medium using chromatographies on DEAE-Cellulose, hydroxyapatite and Sephadex G-100 after salted out with (NH_4)_2SO_4.The enzyme produced levan for sucrose substrate alone, but it produced hetero oligosaccharide when the reaction was made in the presence of hetero oligosaccharide as an acceptor. The mazimum efficiency of transfructosylation from sucrose to acceptor sugar by the enzyme attained about 80% in the case of the acceptor/donor(molar ratio)= 1 or more. Oligosaccharide formed by levansucrase from sucrose and reducing oligosaccharide was non reducing sugar that C2 of fructosyl residue linked to C1 of reducing end of acceptor sugar. Trehalose which is non reducing oligomer, is also able to be an acceptor although the yield of Trehalose-F is very low. In this case, C2 of the fructosyl residue is linked to C6 of glucose residue in trehalose. Levansucrase linked to CH-Sepharose 4B to produce immobilized enzyme which has increased thermal stability (30゜C->40゜C) was used repeatedly to make maltooligosaccharide-F without loss of the activity. Maltooligosaccharide-F synthesized can be used as substrate for differential determination of amylase activity.Other heterooligosaccharides obtained by levansucrase were tested as substrate for determining enzyme specificity and a new -glucosidase in human urine was found. Heterooligosaccharides produced described above are useful as substrate for screening of new carbohydrase.