Thermostability of ancestral mutants of Caldococcus noboribetus isocitrate dehydrogenase |
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Authors: | Iwabata Hisako Watanabe Keiko Ohkuri Takatoshi Yokobori Shin-Ichi Yamagishi Akihiko |
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Institution: | Department of Molecular Biology, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan. |
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Abstract: | We constructed mutant genes of Caldococcus noboribetus isocitrate dehydrogenase containing ancestral amino acid residues that were inferred using the maximal likelihood method and a composite phylogenetic tree of isocitrate dehydrogenase and 3-isopropylmalate dehydrogenase. The mutant genes were expressed in Escherichia coli and the protein products purified. Thermostabilities, reported as the half-inactivation temperatures, for the purified enzymes were determined and compared with that of the wild-type enzyme. Four of the five mutant enzymes have greater thermal stabilities than wild-type isocitrate dehydrogenase. The results are compatible with the hyperthermophilic universal ancestor (commonote) hypothesis. Incorporation of ancestral residues into a modern-day protein sequence can be used to improve protein thermostability. |
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Keywords: | Common ancestor Hyperthermophile Isocitrate dehydrogenase Protein thermostability Caldococcus noboribetus Commonote 3-Isopropylmalate dehydrogenase |
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