Molecular Dynamics Simulations of L-Lactate Dehydrogenase: Conformation of a Mobile Loop and Influence of the Tetrameric Protein Environment

L-lactate dehydrogenase (LDH) catalyses the interconversion of pyruvate and L-lactate in the presence of the coenzyme NADH. Molecular dynamics (MD) simulations have been performed for LDH complexed with NADH and the pyruvate-analogue inhibitor oxamate with the aim of characterising important influences on maintaining the geometry and hydrogen bond network of the active site. Two features in particular were found to dominate. First, the tetrameric protein environment is found to play a significant role in maintaining the active-site geometry. Simulations of the monomer alone reproduce the crystallographic structure poorly, and at least part of the neighboring subunit is necessary to prevent water penetration into the active site and to provide rigidity to the α1G-α2G helix immediately adjacent to the active site. These results offer one explanation for the observation that the monomer is not biologically active. Second, the conformation of Arg109 (part of the mobile loop which closes over the active site) is shown to play a key role in maintaining the active-site geometry. In some simulations, a torsional rotation in the side chain of Arg109 results in the breaking of crystallographic hydrogen bonds which are important for polarising the carbonyl bond of the substrate. This conformational change appears to be a trigger for the opening of the mobile loop. Long-range nonbonded interactions are found to be influential in maintaining the proper crystallographic conformation of Arg109. Thus, we conclude that to adequately model LDH, at least part of the neighboring subunit must be included in the MD simulations and nonbonded interactions must be properly represented to ensure that Arg109 remains in the crystallographic conformation. Out of a set of simulation protocols tested here, one meets both these criteria and will be used for the generation of starting structures for future reaction-mechanism calculations.