バイオサイエンス学科／分子生命科学コース

中村 卓（なかむら・たかし）
Takashi Nakamura

専門分野／生物有機化学、タンパク質工学

職位：講師
学位：博士（工学）（京都大学）

• 京都大学大学院工学研究科博士課程修了
• 独立行政法人理化学研究所・国立環境研究所研究員、山田科学振興財団長期派遣援助研究員（チェコ・マサリク大学）を経て本学へ

研究テーマ

　進化工学的手法による改変酵素の作製とコンピュータ分子モデリングによる分子レベルでの酵素反応機構解析を利用して、実験と計算から高機能な酵素をデザインし、環境汚染物質の分解や産業的に有用な物質の生産に利用していきたいと考えている。

　現在は、以下の3つの酵素について研究を行っている。
（1）農薬やダイオキシン類などによく見られるような有機ハロゲン化合物の一種であるハロ酸を分解するハロ酸脱ハロゲン化酵素L-DEX YL
（2）超好熱性古細菌由来の推定ハロ酸脱ハロゲン化酵素PH0459
（3）超好熱性古細菌由来の耐熱性システイン合成酵素OPSS
　この酵素は医薬品やその前駆物質として極めて有用性の高い非天然アミノ酸を合成することができる。
　L-DEX YLとPH0459は有害物質分解へ、OPSSは有用物質生産への応用を目指している。

Topics of research

Enzymes are molecular machines that catalyze chemical reactions in living organisms. It is of highly scientific interest and practical need to construct enzymes with new catalytic properties and enhanced stability. Tailor-made biocatalysts can be created from wild-type enzymes by protein engineering using computer-aided molecular modeling and site-directed mutagenesis (rational design) or by directed evolution techniques (random mutagenesis).

Rational design usually requires both the availability of the structure of the enzyme and knowledge about the relationships between sequence, structure, mechanism, and function. It is therefore very information-intensive. Using molecular modeling, it has been possible to predict how to increase the selectivity, activity and stability of enzymes.

In contrast, directed evolution involves either a random mutagenesis of the gene encoding the catalyst (e.g. error-prone PCR) or recombination of gene fragments (e.g. DNA shuffling). Libraries thus created are usually assayed using high-throughput technologies to identify improved variants. Recently, combination of computational protein design with directed evolution is often used for creating new enzymes and successful results are obtained in some cases.

The object of my research is to design highly functional enzymes by protein engineering techniques as mentioned above. There are two kinds of enzymes I would like to design. The first enzyme is L-2-haloacid dehalogenase (L-DEX). This enzyme catalyzes the hydrolytic dehalogenation of L-2-haloalkanoic acids to produce the corresponding D-2-hydroxyalkanoic acids. Moreover, PH0459, the haloacid dehalogenase from hyperthermophilic archaeon Pyrococcus horikoshii, is also the target of our project because the enzymes isolated from hyperthermophilic archaeon are usually thermostable as well, and therefore can be used to industrial application. Our goal to the design of L-DEX and PH0459 is to produce the enzyme which can detoxify toxic organic-halogenated compounds.

The second enzyme is O-phosphoserine sulfhydrylase (OPSS). OPSS is found in a hyperthermophilic aerobic archaeon, Aeropyrum permnix K1, is stable at high temperatures as high as 80℃ and produces cysteine more efficiently than other cysteine synthases. It may have potential applications for the efficient production of unnatural amino acids, which are applicable to building blocks for the synthesis of pharmaceuticals and agrochemicals, and the design of new drugs against Trichomonas vaginalis and Mycobecterium tuberculosis. Thus, we are now investigating such a possibility by activity measurement, X-ray crystallography, and molecular modeling.

主な業績論文等

1. Ishikawa, K., Mino, K., Nakamura, T. New Function and Application of the Cysteine Synthase from Archaea . Biochem. Eng. J., 48, 315-322. (2010)
2. Nakamura, T., Yamaguchi, A., Kondo, H., Watanabe, H., Kurihara, T., Esaki, N., Hirono, S., Tanaka, S. Roles of K151 and D180 in L-2-haloacid dehalogenase from Pseudomonas sp. YL: Analysis by molecular dynamics and ab initio fragment molecular orbital calculations. J. Comput. Chem., 30, 2625-2634. (2009)
3. Nakamura, T., Zmock, M., Zdrhal, Z., Chaloupkov, R., Monincov, M., Prokop, Z., Nagata, Y., Damborsk, J. Expression of glycosylated haloalkane dehalogenase LinB in Pichia pastoris. Protein Expr. Purif., 46, 85-91. (2006)