Introduction: Bis-1-chloro-2-[(2-chloroethyl)sulfanyl]ethane, also known as yperite, is a blistering agent and a carcinogen causing nucleotide alkylation. Exposure to yperite leads to major skin, respiratory tract, and eye irritation [1, 2]. Enzymatic degradation of yperite offers many advantages over the traditional methods such as combustion or non-enzymatic chemical degradation. Enzymes can be used to decontaminate materials which would be otherwise destroyed by the chemical degradation, such as military or agricultural equipment [3]. Haloalkane dehalogenases, tested for enzymatic degradation, exhibit low catalytic efficiency, and thus low rate of degradation [3]. Here we describe computational re-design of three of these enzymes towards higher activity with yperite.
Methods: The binding modes of yperite in the active site of selected X-ray structures were obtained using the molecular docking. Subsequently, the minimized systems were analysed by quantum mechanic/molecular mechanic adiabatic mapping along the reaction coordinate of the SN2 reaction. Using this method, transition state conformations were obtained for each system. Using the Rosetta Design [4], we have designed novel enzyme variants, which stabilize the transition state. The binding modes of yperite, thermodynamic parameters of the SN2 substitutions, and thermostability of the novel variants were computationally predicted and compared to the wild-type structures. The relative occurrence of dehalogenation defined by the near attack conformation [5] was obtained by the molecular dynamics.
Results: Using these methods, we obtained 13 new designs which possess thermodynamically feasible mutations, a switch to an exothermic SN2 displacement, much lower activation energy and a higher occurrence of the near attack conformation compared to their corresponding wild-type structures. Selected enzymes will be constructed and characterized experimentally. Novel enzymes re-designed towards higher catalytic activity with yperite could be used for decontamination and bioremediation.