The Structure Ensembles Of Intrinsically Disordered Proteins Studied By Molecular Dynamics Simulations

Intrinsically disordered proteins(IDPs)are a class of proteins that lack a stable conformation and play an important role in many physiological processes in human bodies.IDPs cannot be described in a fixed conformation,but must be represented by a structural ensemble composed of many different conformations.At present,there are many experiments that can detect the structural characteristics of IDPs.However,due to the limitation of experimental conditions and the disordered nature of IDPs,experiments have some limitations in the observation ofIDPs.Molecular dynamics simulation is a method that can investigate the structure and physical/chemical properties of molecules from the atom level,and can well describe the key intermediates and dynamic changes in the structural ensemble of IDPs.At present,molecular dynamics simulation is an important method to study IDPs and is complementary to experimental methods.Molecular dynamics simulation of the structural ensemble of intrinsically disordered proteins is of great significance for studying the function of IDPs.Currently the primary problems faced by MD simulationsis the accuracy of the force field and the efficiency of sampling.Improving sampling efficiency requires the development of enhanced sampling methods,such as replica exchange method and metadynamics methods.In this paper,we use the temperature replica exchange method and the bias exchange metadynamics BE-META method.The main principle of the temperature replica exchange method is that it is easy to cross the energy barrier on the free energy surface at high temperature,while the BE-META method is an enhanced sampling method combining the replica exchange method and the metadynamics method.After the MD simulation is completed,the secondary structure,free energy,chemical shifts and hydrogen bonds of the protein can be analyzed to further understand the structural characteristics of the protein.Firstly,we investigate the regulation mechanism of phosphorylation on the folding of intrinsically disordered P18-R62 fragement of 4E-BP2using temperature replica exchange method.Binding of 4E-BP2 to eIF4E inhibits mRNA cap-dependent translation initiation,while phosphorylation can induce the folding of the P18-R62 fragement of 4E-BP2 from an disordered conformation to a four-stranded parallel?-sheetstructure,with its affinity to eIF4E decreased.Phosphorylated 4E-BP218-62with G37V/G48V mutants cannot fold into ordered structure.In this section we use replica exchange molecular dynamics simulations to study the wild-type WT,phosphorylated wild-type pWT,and phosphorylated mutant pMT of 4E-BP218-62.We have obtained the free energy surfaces(FES)of WT,pWT,and pMT and the main conformations on the FES and found that phosphorylation can significantly increase the proportion of folded conformations in the structure ensemble of 4E-BP218-62.The folding pathway of pWT shows that in the folding process of pWT,two parallel ?/?4,three parallel?1/?4/?3 and four parallel ?1/?4/?3/?2 are formedsuccessively.The ? turn between ?3 and ?4 is formed prior to the ? turn between ?2 and ?3.The formation of two ? turns are the main rate-limiting steps in the folding process.The fraction of contacts and contact maps of WT and pWT show that only strong interactions between pT46 and R20 can promote the folding of pWT,while other long-range interactions between pT37/pT46 and arginines can impede thefolding of pWT.Phosphorylation can cause similar hydrogen bond networks in the two ?turn regions,and local hydrogen bonds may be the primarydriving force for folding of phosphorylated 4E-BP2.The failure of pMT to fold into folded conformations is related to the destruction of the?turnstructures.In addition,we found that phosphorylation can significantly reduce the solvent-accessible surface area of pWT and pMT,which is speculated to be the reason for the decreased affinity of pWT and pMT to eIF4E.This is why pMT,which is also in disordered state,has lower binding affinity to eIF4E than WT.We alsofound that in the disordered WT,the structural ensemble obtained by the CHARMM36m force field is more accurate than the ff99SB-ILDN force field,suggesting that the CHARMM36m force field may be more suitable for the simulation of IDPs than the ff99SB-ILDN force field.We have studied the A?42 using the conventional molecular dynamics method and the bias-exchange metadynamics method.A?42 is an IDP with 42 residues,and its abnormal aggregation is considered to be the key factor in the pathogenesis of Alzheimer's disease.We calculated the chemical shifts,the radius of gyration Rg,the J coupling constant 3 J,and the residual dipole coupling RDC of A?42 after the simulation,and compared them with the experiment determined results.The chemical shifts and 3J show that the results obtained by the BE-META method are closer to the experiment determined results than the conventional MD method,suggesting that BE-META method is more accurate in simulation of the IDPs than conventional MD method.The results of Rg show that,compared with the conventional MD method,the peak of the probability distribution and mean values of Rgfor the BE-META method are closer to the experiment results,and the probability distribution range of the BE-META method is wider,indicating that the results obtained by BE-META are closer to the experiment values and more conformations of different structures can be obtained.The RDC results also show that the figures of the BE-META method is closer to the experiment figures than the conventional MD method.However,the correlation between experimental and calculated RDCs is very low,suggesting that RDCs may not well describe the structure features of IDPs obtained from molecular dynamics simulations.