Martin KlvanaさんはチェコのMasaryk大学から3ヶ月間の予定で 神戸大学大学院自然科学研究科に研究員として滞在され、バイオ レメディエーション酵素の分子設計に関する計算機シミュレーシ ョンの研究に従事されます。9月1日からの赴任にあたり、今までの ご研究のレビューと今後の展望についてお話しいただきますので、 皆様奮ってご参加下さい。

Halogenated aliphatic hydrocarbons are widespread recalcitrant compounds due to massive natural and industrial production. They are environmentally dangerous because of their toxic, genotoxic, teratogenic and irritating effects. Microbial enzymes haloalkane dehalogenases have been studying for decades for their possible use in bioremediation owing to their capability to catalyse hydrolytic dehalogenation of the xenobiotics to harmless product - an alcohol. Practical use of these enzymes requires increase of activity, specificity and thermostability through modification in their structures.

Three haloalkane dehalogenases with experimentally solved three-dimensional structure differ in rate-limiting step of the dehalogenation reaction. The slowest step for Rhodoccocal haloalkane dehalogenase DhaA is release of an alcohol from the active site cavity to bulk solvent. The release may occur through two tunnels called the main tunnel and the slot. Rational computer-assisted re-design of the tunnels could lead to DhaA with higher catalytic activity. Classical molecular dynamics is, however, not appropriate tool for modelling of egress of the product of the reaction because this process is rare event and cannot be observed during nanosecond simulations. We applied Random Accelerated Molecular Dynamics (RAMD) to speed up the egress by applying a force on the product molecule. Using RAMD, we are capable to model export pathways in the time scale from tens to hundreds picoseconds.

RAMD simulations were performed with DhaA in complex with two products of dehalogenation of 1,2,3-trichloropropane - chloride and (S)-2,3-dichloro-1-propanol (S-DCL). Complex was prepared by molecular docking and equilibrated using classical molecular dynamics. Chloride anion solvated by water molecules left the active site through main tunnel after 1735 ps of equilibration phase of molecular dynamics. Its release has never been observed in simulations without alcohol and suggested that chloride leaves the active site with the assistance of water molecules and before S-DCL. RAMD applied on S-DCL showed two pathways for S-DCL, i.e. the main tunnel and the slot, the tunnel being preferred export route.

□　連絡先：田中成典（tanaka2@kobe-u.ac.jp）