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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Acknowledgments
We thank A. M. Grunden of North Carolina State University, USA, for donating the P. furiosus DSM3638 genomic DNA used in the study. This study was supported by the Marine and Extreme Genome Research Center Program, Ministry of Marine Affairs and Fisheries, Republic of Korea.
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Communicated by K. Horikoshi.
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Lee, MH., Yang, SJ., Kim, JW. et al. Characterization of a thermostable cyclodextrin glucanotransferase from Pyrococcus furiosus DSM3638. Extremophiles 11, 537–541 (2007). https://doi.org/10.1007/s00792-007-0061-6
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DOI: https://doi.org/10.1007/s00792-007-0061-6