caa a22 4500 397555830 CHVBK 20180308164801.0 cr unu---uuuuu 161202e199601 xx s 000 0 eng 10.1093/oxfordjournals.jbchem.a021198 doi (NATIONALLICENCE)oxford-10.1093/oxfordjournals.jbchem.a021198 An Aspartate Aminotransferase from an Extremely Thermophilic Bacterium, Thermus thermophilus HB8 [Elektronische Daten] [Akihiro Okamoto, Ryuichi Kato, Ryoji Masui, Akihiko Yamagishi, Tairo Oshima, Seiki Kuramitsu] The aspartate aminotransferase gene (AspAT, EC 2.6.1.1) of an extremely thermophilic bacterium, Thermus thermophilus 11138, was cloned and sequenced, and its gene product was overproduced. The purified T. thermophilus AspAT was stable up to about 80°C at neutral pH. T. thermophilus AspAT was strictly specific for acidic amino acid substrates, such as aspartate, glutamate, and the respective keto acids. The gene coding for T. thermophilus AspAT showed that it comprised 1,155 bp with a high G+C content (70 mol%), and encoded a 385-residue protein with a molecular weight of 42,050. The amino acid sequence of T. thermophilus AspAT deduced from its gene showed about 15, 46, and 29% homology with those from Escherichia coli, Bacillus sp. YM-2, and Sulfolobus solfataricus, respectively. When the amino acid sequence of T. thermophilus AspAT was compared with that of E. coli AspAT, the number of Cys was found to have decreased from 5 to 1, that of Asn from 23 to 9, that of Gln from 16 to 8, and that of Asp from 20 to 13, all of which are known to be relatively labile at high temperatures. Conversely, the number of Pro was increased from 15 to 25, Arg from 22 to 32, and Glu 27 to 37. As shown by the E. coli AspAT structure, there was a marked tendency for the extra prolyl residues to be located around the surface of the molecule. This was quite different from that in the case of RecA protein, which shows an increased number of prolyl residues in the interior of its molecule. Different strategies of different proteins as to prolyl contribution to thermostability have been suggested. Despite the high degree of conservation of active-site residues, Arg292 in E. coli AspAT, which interacts with the distal carboxylate of the substrate, was not found in T. thermophilus AspAT. Arg89 may complement the function of Arg292. © by the Journal of Biochemistry Regular Papers nationallicence aspartate aminotransferase nationallicence gene cloning nationallicence prolyl residue nationallicence thermostability nationallicence Thermus thermophilus nationallicence Okamoto Akihiro Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut Kato Ryuichi Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut Masui Ryoji Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut Yamagishi Akihiko Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-03 aut Oshima Tairo Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-03 aut Kuramitsu Seiki Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut The Journal of Biochemistry Oxford University Press 119/1(1996-01), 135-144 0021-924X 119:1<135 1996 119 jbchem https://doi.org/10.1093/oxfordjournals.jbchem.a021198 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1093/oxfordjournals.jbchem.a021198 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Okamoto Akihiro Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut NATIONALLICENCE 700 1- Kato Ryuichi Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut NATIONALLICENCE 700 1- Masui Ryoji Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut NATIONALLICENCE 700 1- Yamagishi Akihiko Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-03 aut NATIONALLICENCE 700 1- Oshima Tairo Tokyo University of Pharmacy and Life Science, Hachioji, Tokyo 192-03 aut NATIONALLICENCE 700 1- Kuramitsu Seiki Department of Biology, Faculty of Science, Osaka University, Toyonaka, Osaka 560 aut NATIONALLICENCE 773 0- The Journal of Biochemistry Oxford University Press 119/1(1996-01), 135-144 0021-924X 119:1<135 1996 119 jbchem Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford