Department of Life Science and Technology, School of Life Science and Technology, Tokyo Institute of Technology, 4259 Nagatsuta-cho Midori-ku, Yokohama, 226-8501, Japan.
Department of Genome Informatics, Genome Information Research Center, Research Institute of Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka, 565-0871, Japan.
Appl Microbiol Biotechnol. 2019 Dec;103(23-24):9543-9553. doi: 10.1007/s00253-019-10093-w. Epub 2019 Nov 15.
Aliphatic ketones, such as 2-butanone and 3-hexanone, with only one carbon difference among side chains adjacent to the carbonyl carbon are difficult to be reduced enantioselectively. In this study, we utilized an acetophenone reductase from Geotrichum candidum NBRC 4597 (GcAPRD) to reduce challenging aliphatic ketones such as 2-butanone (methyl ethyl ketone) and 3-hexanone (ethyl propyl ketone) to their corresponding (S)-alcohols with 94% ee and > 99% ee, respectively. Through crystallographic structure determination, it was suggested that residue Trp288 limit the size of the small binding pocket. Docking simulations imply that Trp288 plays an important role to form a C-H⋯π interaction for proper orientation of ketones in the pro-S binding pose in order to produce (S)-alcohols. The excellent (S)-enantioselectivity is due to a non-productive pro-R binding pose, consistent with the observation that the (R)-alcohol acts as an inhibitor of (S)-alcohol oxidation.
脂肪族酮，如 2-丁酮和 3-己酮，其羰基碳相邻的侧链之间仅相差一个碳原子，很难进行对映选择性还原。在本研究中，我们利用生香毛霉 NBRC 4597（GcAPRD）中的苯乙酮还原酶将具有挑战性的脂肪族酮，如 2-丁酮（甲基乙基酮）和 3-己酮（乙基丙基酮），分别还原成相应的（S）-醇，对映体过量值（ee 值）分别为 94%和＞99%。通过晶体结构测定，推测残基色氨酸 288 限制了小结合口袋的大小。对接模拟表明，色氨酸 288 对于形成 C-H⋯π 相互作用以正确定向酮在亲核结合构象中起着重要作用，从而产生（S）-醇。优异的（S）-对映选择性归因于非生产性的亲核结合构象，这与（R）-醇作为（S）-醇氧化抑制剂的观察结果一致。
