Molecular cloning and biochemical characterization of two novel oligo‐1,6‐glucosidases from Bacillus mycoides and Thermomyces lanuginosus

Abstract

High-glucose syrup has wide applications in the feed and fermentation industries. However, during its regular production process, because of the slow hydrolysis of a-1,6-glucosidic linkages in maltodextrins by glucoamylase, yields of glucose higher than 96% are rarely achieved. To find a suitable enzyme for the saccharification process, two novel genes encoding oligo-1,6-glucosidases (OGL, EC 3.2.1.10) from Bacillus mycoides (bmogl) and Thermomyces lanuginosus (tlogl) were successfully cloned and overexpressed in Pichia pastoris GS115. At the shake-flask fermentation level, the OGL activities of the two recombinants GS115 (pPIC9K-bmogl) and GS115 (pPIC9K-tlogl) were 994 and 1219 U/mL, respectively; and mature enzymes around 66-68 kDa were purified for characterization. Recombinant enzyme TlOGL exhibited higher thermostability than ever reported for OGLs, whereas BmOGL was stable under acidic pH, ranging from 4.0 to 8.0. Among the substrates tested, these two recombinant enzymes efficiently hydrolyzed isomaltose and isomaltotriose, but had no activity against maltose and maltotriose. Besides these characteristics, nearly complete hydrolysis of isomaltotriose and 90% of isomaltooligosaccharides (IMOs) into glucose was also observed for them, which makes them good candidates for subsequent use in improving the yield of glucose from starch. To the best of our knowledge, this is the first report on the expression and characterization of OGLs from B. mycoides and T. lanuginosus.

Pullulanase with high temperature and low pH optima improved starch saccharification efficiency

“…Since the commercially available pullulanase has no activity on isomalto-oligosaccharides 36 , optimization of saccharification endpoint and using optimized formula of glucoamylase blend may further improve the maximal DX value. Besides, other enzymes that can hydrolyze isomalto-oligosaccharides 37 may involve in glucoamylase blend preparation.…”