Synthesis of improved long-chain isomaltooligosaccharide, using a novel glucosyltransferase derived from Thermoanaerobacter thermocopriae, with maltodextrin

TG, transglucosidase; TtTG, T. thermocopriae-derived TG; G, glucose; IG2, isomaltose; IG3, isomaltotriose; P, panose; IG4, isomaltotetraose; IG5, isomaltopentaose; IMO, isomaltooligosaccharides; L-IMO, long-chain IMO; ALO, anomalously linked oligosaccharides; DP, degree of polymerization; o, glucose; Φ, reducing end; ̶, α-1,4 linkage; ┓, α-1,6 linkage. [Display omitted] •Identified novel Thermoanaerobacter thermocopriae-derived transglucosidase (TtTG).•TtTG showed optimal pH and temperature of 4.0 and 60 °C, respectively.•TtTG produced high polymerization degree-isomaltooligosaccharides using maltodextrin.•The α-1,6/α-1,4 linkage ratio of isomaltooligosaccharides produced by TtTG was three times higher. Isomaltooligosaccharide (IMO), considered to be a prebiotic, reportedly has health effects, particularly in terms of digestion; however, the prebiotic effects of IMOs depend largely on the degree of polymerization. Currently, IMOs are commercially produced using transglucosidase (TG) derived from Aspergillus niger. Here, we report a novel Thermoanaerobacter thermocopriae-derived TG (TtTG) that can produce long-chain IMOs (L-IMOs) using maltodextrin as the main substrate. A putative carbohydrate-binding gene comprising carbohydrate-binding module 35 and glycoside hydrolase family 15 domain was cloned and successfully overexpressed in Escherichia coli BL21 (DE3) cells. The resulting purified recombinant enzyme (TtTG) had a molecular mass of 94 kDa. TtTG displayed an optimal pH of 4.0 (higher than that of commercial TG) and an optimal temperature of 60 °C (same as that of commercial TG). TtTG also enabled the synthesis of oligosaccharides using various saccharides, such as palatinose, kojibiose, sophorose, maltose, cellobiose, isomaltose, gentiobiose, and trehalose, which acted as specific acceptors. TtTG could also produce a medium-sized L-IMO, different from that by dextran-dextrinase and TG, from maltodextrin, as the sole substrate. Thus, the novel combination of maltodextrin and TtTG shows potential as an effective method for commercially producing L-IMOs with improved prebiotic effects.