QM/MM MD and free energy simulations of G9a-like protein (GLP) and its mutants: understanding the factors that determine the product specificity.

Yuzhuo Chu,Hong Guo,Jianzhuang Yao

doi:10.1371/journal.pone.0037674

Abstract

Certain lysine residues on histone tails could be methylated by protein lysine methyltransferases (PKMTs) using S-adenosyl-L-methionine (AdoMet) as the methyl donor. Since the methylation states of the target lysines play a fundamental role in the regulation of chromatin structure and gene expression, it is important to study the property of PKMTs that allows a specific number of methyl groups (one, two or three) to be added (termed as product specificity). It has been shown that the product specificity of PKMTs may be controlled in part by the existence of specific residues at the active site. One of the best examples is a Phe/Tyr switch found in many PKMTs. Here quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) and free energy simulations are performed on wild type G9a-like protein (GLP) and its F1209Y and Y1124F mutants for understanding the energetic origin of the product specificity and the reasons for the change of product specificity as a result of single-residue mutations at the Phe/Tyr switch as well as other positions. The free energy barriers of the methyl transfer processes calculated from our simulations are consistent with experimental data, supporting the suggestion that the relative free energy barriers may determine, at least in part, the product specificity of PKMTs. The changes of the free energy barriers as a result of the mutations are also discussed based on the structural information obtained from the simulations. The results suggest that the space and active-site interactions around the ε-amino group of the target lysine available for methyl addition appear to among the key structural factors in controlling the product specificity and activity of PKMTs.

Highlights

The tails of histone proteins are subject to a variety of posttranslational modifications, and these modifications are believed to be a part of the histone code for chromatin regulation [1]
PKMTs may be classified based on their ability to transfer one, two or three methyl groups from S-adenosyl-L-methionine (AdoMet, the methyl donor) to the e-amino group of target lysine, a special property of the enzymes that is termed as product specificity [1,4]
The dynamic information of reactant complexes gathered from the quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) simulations were examined, and the results suggest that the ability of the reactant complex to form the reactive configuration may be used as an important indicator for the product specificity of PKMTs

Summary

Introduction

The tails of histone proteins are subject to a variety of posttranslational modifications, and these modifications are believed to be a part of the histone code for chromatin regulation [1]. For H3K9 G9a methyltransferase, the FRY mutant at the Phe/Tyr switch site (F1152 in G9a) was found to alter the enzyme from a dimethyltransferase to a mono-methyltransferase [14]; the equivalent residue in its closely relative G9a-like-protein (GLP) is F1209. Previous studies on SET7/9, SET8 and DIM-5 [13,17,18,19] have suggested that this Phe/Tyr switch may affect product specificity through altering the affinity of an active site water molecule, as the dissociation of this water molecule is likely to make it easier for further methyl addition [13,17,18,19]. It would be of interest to examine whether this is the case for GLP or G9a as well

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Conclusion