Enhancement of thermostability of kojibiose phosphorylase from Thermoanaerobacter brockii ATCC35047 by random mutagenesis |
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Authors: | Yamamoto Takuo Mukai Kazuhisa Yamashita Hiroshi Kubota Michio Fukuda Shigeharu Kurimoto Masashi Tsujisaka Yoshio |
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Affiliation: | Amase Institute, Hayashibara Biochemical Laboratories, Inc., 7-7 Amase Minami-machi, Okayama 700-0834, Japan. amaseken@hayashibara.co.jp |
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Abstract: | Random mutation by error-prone PCR was introduced into kojibiose phosphorylase from Thermoanaerobacter brockii ATCC35047. One thermostable mutant enzyme, D513N, was isolated. The D513N mutant enzyme showed an optimum temperature of 67.5-70 degrees C (the wild type, 65 degrees C), and thermostability up to 67.5 degrees C (the wild type, up to 60 degrees C). The half-lives of D513N were estimated to be 135 h at 60 degrees C, 110 min at 70 degrees C and 6 min at 75 degrees C, respectively. They were about 1.6-fold, 7-fold and 6-fold longer than those of the wild-type enzyme, respectively. |
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Keywords: | kojibiose phosphorylase kojioligosaccharide random mutation thermostability |
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