Molecular Dynamics of Electrostatic Channeling on the Surface of Malate Dehydrogenase-Citrate Synthase

Abstract

Multi-enzyme catalytic cascade reactions exist in many metabolic pathways in nature; the benefits of these reactions include high efficiency and reduced time and labor costs [1].The design and application of efficient cascade reactions in biofuel cells, sensors, and chemical conversion requires an understanding of intermediate channeling between adjacent cascade active sites. Electrostatic channeling, for example, utilizes electrostatic force to control the direct transfer of charged intermediates [2]. Here, we study the electrostatic channeling of an oxaloacetate (OAA) intermediate between malate dehydrogenase (MDH) and citrate synthase (CS) active sites within the tricarboxylic acid (TCA) cycle.The MDH-CS supramolecular assembly includes a positively charged surface between neighboring active sites that guides the movement of the negatively charged oxaloacetate, thus improving the transfer efficiency [3].To study the interaction between OAA and the charged enzyme surface and identify OAA’s transfer pathways, we simulated this system using molecular dynamics (MD). Specifically, since both MDH and CS have two identical subunits and thus two active sites, we ran 50 MD simulations for 10 ns near each of the four active sites, resulting in 200 parallel runs of simulations. Then we analyzed the resulting MD dataset with a Markov State Model (MSM) [4].We identified multiple energy basins in MSM, including four active site basins, a desorption basin, and an intermediate basin. MSM successfully predicted the the transition rates between those energy basins and identified the dominant transport pathways of OAA from MDH to CS. The visualization demonstrates that the dominant transport pathway crosses Arg65, providing further support for the experimental observation that Arg65 plays an critical role in the electrostatic transport process of OAA [3].This study improves the field’s understanding of natural substrate channeling mechanisms and can help with the design of artificial cascade reactions. References K. C. Nicolaou, D. J. Edmonds, and P. G. Bulger, Angew. Chemie - Int. Ed., 45, 7134–7186 (2006). doi:10.1002/anie.200601872. I. Wheeldon, S. D. Minteer, S. Banta, S. Calabrese Barton, P. Atanassov, and M. Sigman, Nat. Chem (2016). doi:10.1038/NCHEM.2459 B. Bulutoglu, K. E. Garcia, F. Wu, S. D. Minteer, and S. Banta, ACS Chem. Biol., 11, 2847–2853 (2016). doi:10.1021/acschembio.6b00523F. Noé, C. Schütte, E. Vanden-Eijnden, L. Reich, and T. R. Weikl, Proc. Natl. Acad. Sci. U. S. A., 106, 19011–19016 (2009). doi:10.1073/pnas.0905466106 Figure 1