The Effect Of L210F Mutation On The CRAC Ion Channel Studied By Molecular Dynamics Simulations

Calcium ion,as an essential second messenger in cells,regulates a wide range of physiological processes,such as gene transcription,cell proliferation,differentiation and apoptosis.Calcium ion channels are the basis for ion permeation and regulation across and within cells.The calcium-induced calcium influx is one of the main Ca2+regulation mechanisms,and the CRAC channel is the primary channel to mediate calcium influx.Although the passage of calcium ions through CRAC channels into cells has been known for decades,its molecular structure and function have not been studied at the atomistic detail until recently.It has been observed experimentally that the L210F mutation of the CRAC channel can activate the channel without the involvement of the activator.This paper will take L21 OF mutation as the breakthrough point of the opening mechanism of CRAC channel for further in-depth exploration.Based on the existing research showing that the L210F mutation may lead to an open state CRAC,we studied the impact of this single non-pore residue mutation on the conformation of the CRAC channel by using molecular dynamics(MD)simulations.We used the MD package Gromacs to simulate the conformational changes induced by the L210F mutation.The trajectories are then analyzed with the MDAnalysis package and the built-in analysis program in Gromacs.In particular,we investigated the following:1.The effect of L210F mutation on the conformation of the CRAC ion channel,both globally and locally.In this paper,the wild-type and mutant structures were simulated in parallel.By comparison,the results showed that the radius of the basic region and the filter region underwent an appreciable expansion in the mutant simulations.Further analysis showed that after the L210F mutation,the residue 210 had a greater degree of freedom and a larger rotation angle than that of the wild type,which in turn reduced its restriction on TM1 and enhanced its interaction with the adjacent subunit TM3.2.The effect of the L210F mutation on the surrounding environment of the CRAC channel.Our analysis of ion distribution around the channel showed that L210F significantly increased the number of water molecules between the filter and hydrophobic regions of the channel,and the increase of pore hydration would facilitate the passage of ions,which was consistent with the experimental result that L210F led to spontaneous activation of the channel.We did not observe a significant difference in the distribution of Cl-in the two types of simulations,but found that the penetration of cationic Na+in the channel was deeper in the mutant,further indicating that the mutation L210F may lead to an open state of the CRAC channel.We have not observed complete permeation of Na+through the channel,which may be associated with the low conductivity of CRAC,as well as the simulation timescale not long enough.In summary,the L210F mutant may spontaneously evolve into the open state,as evidenced by our simulation results.The increase of the pore radius and pore hydration can reduce the energy barrier of Ca2+ passing through the channel,thus allowing ions to permeate.The simulation results in this paper are consistent with the previous experimental study of the L210F mutant,and provide atomistic details as a plausible explanation.