A significant role of Arg41 residue in the enzymatic reaction of haloacid dehalogenase l-DEX YL studied by QM/MM method

Theoretical prediction and experimental verification on enantioselectivity of haloacid dehalogenase l-DEX YL with chloropropionate

“…The ONIOM calculations were performed with the PM3 [27] method for the QM region and the Amber 96 force field [28] for the MM region (PM3:Amber) using the Gaussian09. It is noted that the use of PM3 method in the present study was validated in the earlier work [15] through comparison with the density-functional-theory calculations. One hundred fiftyfive atoms including the side chains of all residues of the QM region were then structurally optimized, while the other atoms were fixed during the optimization.…”

“…A structure of the enzyme-substrate complex of l-DEX YL and l-2-CPA in our previous study [15] was used as a reference structure designated as WT/L for comparison. Structures of enzyme-substrate complex of l-DEX YL and d-2-CPA were constructed by using GaussView5 [23].…”

“…The distance between nucleophilic oxygen atom closer to the d-2-CPA of the aspartic acid (Asp10) and the asymmetric carbon atom of the d-2-CPA was selected as an internal reaction coordinate. The structure in the QM region was fully optimized with the exception of the driving coordinate, where ␣-carbons of the twelve residues in the QM region were fixed [15]. Then a TS structure was obtained from the maximum-point of the potential energy curve, and a reactant structure (enzyme-substrate complex) and a product structure (ester intermediate) were determined with the intrinsic reaction coordinate method [29].…”

“…strain YL cell culture in a medium containing 2-chloropropionate (2-CPA) as the sole carbon source [5]. The reaction mechanism of l-DEX YL has been analyzed by site-directed mutagenesis [6], X-ray crystallography [7][8][9], mass spectrometry [10][11][12], 18 O-incorporation experiments [13], and computer simulations with molecular dynamics (MD), fragment molecular orbital (FMO) [14] and quantum-mechanical/molecularmechanical (QM/MM) methods [15]. Site-directed mutagenesis experiments have revealed that some amino acid residues are essential for the catalytic function of l-DEX YL [6].…”

“…The contribution of each amino acid residue in l-DEX YL was examined through computer simulations with MD and FMO methods [14]. Recently, QM/MM simulations determined the transition state (TS) structure of the enzyme-substrate complex and elucidated its catalytic mechanism in the reaction of ester intermediate formation at the molecular level [15].…”