岡山大学農学部 Acta Medica Okayama 0474-0254　 97 1 2008 海産性好熱性細菌 Rhodothermus marinus 由来イソアミラーゼ の精製，性質検討及びＸ線結晶構造解析 1 7 EN Akiko Tachibana Takashi Tamura Katsumi Imada Miki Kinoshita Keiichi Namba Noriko Tsutsumi Miyoko Hashida Hiromichi Sakaguchi Kenji Inagaki The isoamylase gene from Rhodothermus marinus was cloned into and expressed in Escherichia coli Top 10. As a result of characterization of purified R. marinus isoamylase. the enzyme had an optimum pH of 4.0 and optimum temperature of 70℃. Thermal inactivation studies of the purified R. marinus isoamylase revealed the enzymatic activity to be uninfluenced after one hour incubation at 60℃. These results suggest that R. marinus isoamylase has high thermostability. The crystallization and crystal structure analysis of R. marinus isoamylase was performed. The three-dimensional structure at 1.9Å resolution was determined in complex with the panose. R. marinus isoamylase is composed of three domains N, A and C, and, has a (β/α)8-barrel in domain A. The secondary structural alignments of the R. marinus isoamylase and P. amyloderamosa isoamylase was carried out. They have the four active-site consensus regions characteristic of the α-amylase family. And the essential residue of the α-amylase family (D359, E395, and D467) was conserved in these enzymes. R. marinus isoamylase has shorter loops than P. amyloderamosa isoamylase. And R. marinus isoamylase had no Ca2+ binding site. These results are thought to be factors of thermostability of R. marinus isoamylase. No potential conflict of interest relevant to this article was reported.