Molecular cloning and characterization of trehalose synthase from Thermotoga maritima DSM3109: Syntheses of trehalose disaccharide analogues and NDP-glucoses

Soo In Ryu, Jeong Eun Kim, Nguyen Thi Huong, Eui Jeon Woo, Sung Kwon Moon, Soo Bok Lee

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

A gene (ORF TM0392) encoding a putative trehalose synthase (TmTreT) in Thermotoga maritima was cloned and expressed in Escherichia coli. The recombinant enzyme was purified to homogeneity by heat treatment and a glutathione-sepharose affinity column chromatography. The purified enzyme existed exclusively as a monomer in a native state. The optimum pH and temperature for this enzyme were 6.0 and 65°C. The glutathione-S-transferase (GST)-fusion enzyme had greater thermostability than thrombin-treated free enzyme. TmTreT had diverse substrate specificities. The enzyme effectively created a free trehalose from several nucleoside diphosphate (NDP)-glucoses as a donor and glucose as an acceptor. Inversely, the enzyme was also capable of employing several NDPs such as UDP, ADP, GDP, and CDP with trehalose to produce corresponding NDP-glucoses. The enzyme was able to employ other monosaccharides, such as mannose and fructose, as acceptors to synthesize disaccharide analogues of trehalose. The mannose-containing analogue was not hydrolyzed by trehalase and the rat intestinal enzymes. Furthermore, the analogue showed a competitive inhibition to the intestinal disaccharidases with Ki values of approximately 0.8-1.6mM. The results suggest that the enzyme is an useful trehalose synthase that can regenerate NDP-glucoses from NDPs and produce the inhibitory trehalose analogues of indigestible disaccharides.

Original languageEnglish
Pages (from-to)249-256
Number of pages8
JournalEnzyme and Microbial Technology
Volume47
Issue number6
DOIs
Publication statusPublished - 2010 Nov 8

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Biochemistry
  • Applied Microbiology and Biotechnology

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