• KSBB Journal
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• 제14권4호
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• pp.429-433
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• 1999

$\beta$-Glucosidaes from Cellvibrio gilvus(CG) was successfully overproduced in soluble form in E. coli with the coexpression of GroEL/ES/. Without the GroEL/ES protein, the $\beta$-glucosidase overexpressed in E. coli constituted a huge amount(80%) of total cellular protein, but was localized in the insoluble fraction, and little activity was detected in the soluble fraction. Coexpression of the E. coli GroEL/ES had a drastic impact on the proper folding of the $\beta$-glucosidase; 20% of the overexpressed enzyme was recovered in the soluble fraction in active form. Similar effects of GroEL/ES were also observed on the overexpressed $\beta$-glucosidase from Agrobacterium tumefaciens(AT). And pET28(a)-RGRAR, partially deleted mutant lacking 5-amino acid residues at carboxy teminus also could be folded into an active form when expressed with the molecular chaperonin GroEL/ES, and its activity was higher than that of the without GroEL/ES system, In addition, the synergistic effect of GroEL/ES and the low induction temperature were important factors for solubilization of the inclusion body from overproduced $\beta$-glucosidases.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2001년도 Proceedings of 2001 International Symposium
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• pp.179-182
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• 2001

Two intracellular enzymes, cellobiose phosphorylase (CBP) and cellodextrin phosphorylase (CDP) are involved in the phosphorolytic pathway in cellulose degradation. Those enzymes are considered to be useful in syntheses of oligosaccharides because the reactions are reversible. CBP from Cellvibrio gilvus and CDP from Clostridium thermocellum YM-4 were cloned and over-expressed in Escharichia coli. Both the enzyme reactions showed ordered bi bi mechanism. Acceptor specificity of CBP in the reverse reaction was determined. Several $\beta$-l,4-glucosyl disaccharides were synthesized by using the reaction. A new substrate inhibition pattern, competitive substrate inhibition, was also found in the reverse reaction of CBP Cellobiose was produced from sucrose at a high yield by a combined action of three enzymes including CBP