Molecular Dynamics Simulation Of SETD3 Mediated Histidine Methylation On ?-actin

The N?₂-methylation of H73 in?-actin has been known for more than 50 years,and this methylation plays an important role in controlling cellular functions of actin filaments and the release pathway of phosphate groups.The biochemical experiments showed that the enzyme for the His73 methylation of?-actin is SETD3.In the last few years,the crystal structures of the SETD3 complexes have been determined.These crystal structures are of great importance for studying the mechanism of histidine methylation catalyzed by SETD3 for which the details of the catalytic process are still not clear.The combination of quantum mechanical/molecular mechanical(QM/MM)method and molecular dynamics(MD)simulations play an important role in the study of enzyme-substrate catalytic reaction mechanism in biological system.Here we use these computational approaches to examine the structures of reaction and transition states and understand dynamic properties of the SETD3 complex in the process of methylation.We also study the catalytic mechanism of SETD3.MD and free energy simulations based on QM/MM potential are performed for SETD3 wild type and its N255A and Y312F mutants respectively.The results demonstrate that the reactive state for methylation in which H73 adopts the N_?1-H?tautomeric form is stable and the H73 is well positioned at the SETD3 active site for accepting the methyl group from S-adenosyl-L-methionine(SAM).Moreover,the imidazole ring of H73 in the reactive state has a similar orientation to methylated H73(H73Me)in the crystal structure of product complex.Thus,the imidazole ring of H73 does not undergo rotation during the methyl transfer,and this suggestion is further supported by the results of the PMF simulations for the rotation around the C?-C?bond in the reactive state as well as classical MD simulations.The free energy simulations are also performed for the methyl transfer from SAM to N?2 of the N?1-H?tautomer of H73 in wild-type SETD3 as well as in N255A and Y312F mutants.It is shown that the free energy barrier increases by approximately1.5-3 kcal·mol^-1 as a result of these mutations,consistent with the experimental observations.The implication for the existence of the stable reactive state for methylation with well-positioned H73 in the SETD3-substrate complex is discussed,and the origin of the increase of the free energy barrier as a result of the mutations is proposed.The study presented here expands our understanding of the catalytic mechanism of SETD3,especially provides important information to further understand the mechanism of the histidine methylation.