Characterization of a thermostable cyclodextrin glucanotransferase from <Emphasis Type="Italic">Pyrococcus furiosus</Emphasis> DSM3638 |
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Authors: | Myoung-Hee Lee Sung-Jae Yang Jung-Woo Kim Hee-Seob Lee Jung-Wan Kim Kwan-Hwa Park |
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Institution: | (1) Center for Agricultural Biomaterials, and Department of Food Science and Biotechnology, School of Agricultural Biotechnology, Seoul National University, Seoul, 151-921, South Korea;(2) Department of Biology, University of Incheon, Incheon, 402-749, South Korea |
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Abstract: | A gene that encodes the enzyme Pyrococcus furiosus cyclodextrin glucanotransferase (PFCGT) was cloned in Escherichia
coli. PFCGT was highly expressed in recombinant E. coli after compensation for codon usage bias using the pRARE plasmid. Purified PFCGT was extremely thermostable with an optimal
temperature and pH of 95°C and 5.0, respectively, retaining 97% of its activity at 100°C. Incubation at 60°C for 20 min during
the purification process led to a 1.5-fold increase in enzymatic activity. A time course assay of the PFCGT reaction with
starch indicated that cyclic α-1,4-glucans with DPs greater than 20 were produced at the beginning of the incubation followed
by an increase in β-CD. The major final product of PFCGT cyclization was β-CD, and thus the enzyme is a β-CGTase. |
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Keywords: | Hyperthermophile Pyrococcus furiosus Pyrococcus furiosus cyclodextrin glucanotransferase (PFCGT) |
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