Sunday, April 29, 2007

Synthesis of long isomaltooligosaccharides using sucrose

Xing-ji Jin1, Eun-Ah Ko2, Mi-Young Seo3, Nahyun M. Kim4, and Doman Kim2. (1) Department of Fine Chemical Engineering, Jeonnam National University, 300 Yongbong-dong, Gwangju, 500-757, South Korea, (2) School of Biological Sciences and Technology, Jeonnam National University, 300 Yongbong-dong, Gwangju, 500-757, South Korea, (3) Molecular Bioprocess Research Center, KRIBB, Jeonbuk, 580-185, South Korea, (4) Kwangju Foreign School, Yangsan-dong, Gwangju, 500-220, South Korea

Isomaltooligosaccharides and oligodextrans appear to harbor a great deal of promise as prebiotics, and have also been employed in treatments for chronic constipation and hyperlipidemia. The most frequently applied enzymatic method of IMO production involves amylases, (neo) pullulanase and glucosidase acting on starch. Another method involves the conversion of a mixture of sucrose and glucose to dextransucrase via an acceptor reaction. The other primary method involves the use of immobilized dextransucrase and soluble dextranase in sucrose solution. Each process requires improvements with regard to the synthesis of oligosaccharides, including the co-synthesis of dextran as a side product and/or the amount of un-reacted acceptor remaining after the acceptor reaction digestion steps, as well as the synthesis of branched isomaltooligosaccharides rather than linear isomaltooligosaccharides in dextran-hydrolysis by dextranase.
A hybrid gene (dxsr) possessing both dextranase and dextransucrase activities was constructed for long isomaltooligosaccharide (LIMO) synthesis using sucrose. For the dextranase gene, Arthrobacter oxydans was isolated and the biochemical properties of its dextranase were evaluated. The endo-dextranase gene (dex2) was cloned and expressed in Escherichia coli. The specific activity of the purified enzyme was 2.59 U/mg. The dextransucrase gene (dsrBCB4) was also isolated from Leuconostoc mesenteroides B-1299CB4, after which the DXSR hybrid enzyme was constructed. It consists of 2,099 amino acids, with a calculated molecular mass of 234.3 kDa. The DXSR was utilized to synthesize LIMO in high yield using only sucrose with various concentrations. The optimum reaction condition for LIMO and the function of LIMO as an anti-cariogenic material were studied.