MOLECULAR DYNAMIC MODELLING OF A NOVEL PLCG2 VARIANT REVEALS KEY PROTEIN STRUCTURAL DIFFERENCES

Abstract

A novel rare coding variant, associated with Alzheimer's disease (AD), in PLCG2 (rs72824905/p.P522R, P=5.38x10-10), was recently identified using whole-exome-microarray [1]. PLCG2 contains a TIM-barrel, where the active site, catalytic residues and a Ca2+ binding site are found. The P522R mutation can be found in the sPH domain which, structurally, is near the TIM-barrel. This domain is a part of the auto-inhibitory component of the PLCG2 gene, and a mutation here can impact upon PLCG2's activation. We hypothesize that this protective variant alters the function of the protein, via a shift in the position of the sPH loop caused by the change from a neutral-to-positive amino acid. Homology modelling was used to construct PLCG2, this was carried out using I-TASSER. A model with a low C-score, a measure of protein quality, was selected and further scrutinised using PROCHECK. The wild type and mutated proteins were subjected to 50ns of Molecular Dynamics (MD). MD was carried out, in triplicate, using GROMACS. All simulations were all run in the NPT ensemble, with periodic boundary conditions, at a temperature of 310K. Resulting structures were analysed using RMSD and visualised for structural differences using VMD. MD results suggest a structural change to the protein, caused by the P522R mutation, which impacts upon the availability of the active site for binding. RMSD results show no statistically significant changes to the overall flexibility of the protein. This, suggests a localised mutational effect. The arginine amino acid, carrying a positive charge, is attracted to the surrounding negatively charged amino acids causing a shift in the sPH loop resulting in the partial covering of the entrance of the active site. This protective rare variant may cause a large functional change to the PLCG2 protein, by blocking the active site, something that we will test in a wet lab environment. PLCG2 has very little known association with AD and the impact of this novel coding change could prove to further enhance our knowledge into the vast genetic landscape of the disease. 1. Sims R et al,. Rare coding variants in PLCG2, ABI3 and TREM2 implicate microglial-mediated innate immunity in Alzheimer's disease. Nature Genetics (2017), in press.