Analysis of dextrin dextranase from Gluconobacter oxydans

[摘要] Dextran is a high value glucose polymer used in medicine and an array of laboratorytechniques. It is synthesised by lactic-acid bacteria from sucrose but has also reportedlybeen produced by Gluconobacter oxydans (G. oxydans) from a range ofmaltooligosaccharides (MOS) via the action of dextrin dextranase (DDase). In thisstudy the presence of DDase is investigated in two G. oxydans strains (ATCC 621Hand ATCC 19357) and shown to be present in the ATCC 19357 strain, but not in theATCC 621H strain. The enzyme was partially purified from the ATCC 19357 strain,and its kinetic properties investigated. The partially purified protein was also digestedwith trypsin, and de novo peptide sequences obtained from it. Several attempts weremade to obtain the gene coding for the DDase. These include amplifying an openreading frame from the G. oxydans genome coding for a glycosyltransferase with theapproximate molecular weight of the DDase, using the peptide sequences obtainedfrom the partially purified protein to design degenerate PCR primers and the productionof a genomic DNA library for functional screening in E. coli. None of these approachesled to the successful isolation of the extracellular DDase sequence.

[发布机构] Stellenbosch University