Carboxy-Terminal Region of a Thermostable CITase from Thermoanaerobacter thermocopriae Has the Ability to Produce Long Isomaltooligosaccharides

• Published in: Journal of microbiology and biotechnology Vol. 29; no. 12; pp. 1938 - 1946
• Main Authors: Jeong, Woo Soo, Lee, Yu-Ri, Hong, Seong-Jin, Choi, Su-Jeong, Choi, Ji-Ho, Park, Shin-Young, Woo, Eui-Jeon, Kim, Young Min, Park, Bo-Ram
• Format: Journal Article
• Language: English
• Published: Korea (South) 한국미생물·생명공학회 28-12-2019
• Subjects: Catalytic Domain; Cloning, Molecular; Enzyme Stability; Escherichia coli - genetics; Glucosyltransferases - chemistry; Glucosyltransferases - genetics; Glucosyltransferases - metabolism; Glycoside Hydrolases; Hydrogen-Ion Concentration; Molecular Weight; Oligosaccharides - metabolism; Polymerization; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Sequence Alignment; Sequence Analysis, Protein; Temperature; Thermoanaerobacter - enzymology; Thermoanaerobacter - genetics; 생물학; long isomaltooligosaccharides; Cycloisomaltooligosaccharide glucanotransferase; Thermoanaerobacter thermocopriae; cyclodextran
• ISSN: 1017-7825; 1738-8872
• DOI: 10.4014/jmb.1910.10022

Isomaltooligosaccharides (IMOs) have good prebiotic effects, and long IMOs (LIMOs) with a degree of polymerization (DP) of 7 or above show improved effects. However, they are not yet commercially available, and require costly enzymes and processes for production. The Nterminal region of the thermostable
cycloisomaltooligosaccharide glucanotransferase (TtCITase) shows cyclic isomaltooligosaccharide (CI)-producing activity owing to a catalytic domain of glycoside hydrolase (GH) family 66 and carbohydrate-binding module (CBM) 35. In the present study, we elucidated the activity of the C-terminal region of TtCITase (TtCITase-C; Met740-Phe1,559), including a CBM35-like region and the GH family 15 domain. The domain was successfully cloned, expressed, and purified as a single protein with a molecular mass of 115 kDa. TtCITase-C exhibited optimal activity at 40°C and pH 5.5, and retained 100% activity at pH 5.5 after 18-h incubation. TtCITase-C synthesized α-1,6 glucosyl products with over seven degrees of polymerization (DP) by an α-1,6 glucosyl transfer reaction from maltopentaose, isomaltopentaose, or commercialized maltodextrins as substrates. These results indicate that TtCITase-C could be used for the production of α-1,6 glucosyl oligosaccharides with over DP7 (LIMOs) in a more cost-effective manner, without requiring cyclodextran.