Effect Of The R126C Mutation On The Structure And Function Of The Glucose Transporter GLUT1:A Molecular Dynamics Simulation Study

Background and objectiveGlucose transport protein(Glut)is responsible for basal glucose uptake and is expressed in most tissues,and is the most basic requirement for cells to obtain glucose.Glucose transporter 1 deficiency syndrome(GLUT1DS)is caused by haploinsufficiency of SLC2A1 encoding Glut-1,resulting in impaired transport of hexose into the brain.Studies have shown that Glut-1 is not only involved in the normal biological function and metabolism of cells,but also highly expressed in various cancer tissues,including lung cancer,thyroid cancer,breast cancer,etc.The expression level of Glut-1 is strongly associated with the histological type of tumor cells,the differentiation of lymph node metastasis,as well as disease progress and prognosis.In addition,the large demand of tumor cells for glucose also makes Glut-1 a molecular marker for tumor diagnosis.Glut-1 mutation(as exemplified by R126C)can lead to premature absence epilepsy and myoclonic dystonia syndrome(MDS),in which MDS can be fatal.In the present study,the effect of R126C mutation in MDS on structural stability and substrate transport of Glut-1 was investigated.Our study provides insights for understanding the role of glucose transporter,developing drugs for glucose metabolism disorders and the diagnosis of tumors.Methods1.UniProtKB,UniRef,DUET,MUpro,I-Mutant2.0,iStable bioinformatics tools were used to predict free energy after Glut-1 mutation;PredyFlexy and VarSite servers were used to predict the effects of R126C mutation on the flexibility of Glut-1 protein structure.2.Molecular dynamics(MD)simulations were used to further characterize the effect of the R126C mutation on the structural stability of Glut-1.Based on the constructed simulation system,three independent 500ns MD simulations were performed systematically.The protein structural,volatility,adaptive guided molecular dynamics,principal component,secondary structure,porcupine plot,dynamic crosscorrelation plot analysis and free energy analysis of glucose through Glut1 channels were analyzed during each simulation.Results1.After the Glut-1 mutation calculated by the bioinformatics tools DUET,mCSM,SDM,MUpro,I-Mutant2.0 and iStable,the free energy(ΔΔG)decreased by 2.405 kcal/mol,2.193 kcal/mol,1.500 kcal/mol,0.8995kcal/mol,1.310kcal/mol and 0.685kcal/mol,respectively.2.The R126C mutation resulted in greater conformational heterogeneity than wild-type Glut-1 as revealed by the results from RMSD,RMSF and Rg.There was fewer intramolecular hydrogen bonds observed in Glut1 with R126C mutation compared to the wild-type Glut-1 shown by hydrogen bonds analysis.The principal component analysis further showed that R126C mutation can alter the local conformational structure.secondary structure analysis confirms that the R126C mutation has a distal effect on the structure of Glut1,whereas the stability within the position 126 remains unchanged.Both the graph analysis and the porcupine graph analyze the high-intensity flexible motion of the mutants;the free energy analysis of glucose through the Glut1 channel shows that the R126C mutation reduces the PMF value of glucose through Glut-1,and the free energy of glucose transport through Glut1 is significantly The increase,especially at the mutated site,suggests that the substrate is not stable on the cytoplasmic side of the channel,and the passage of glucose through the channel is hindered.ConclusionIn the present study,the R126C mutation was predicted to reduce the structural stability of Glut-1 based on a series of bioinformatics analysis.We found that the nonsynonymous mutation Arg126Cys(R126C)changes the structural characteristics of the functionally important domain of the glucose transporter,These results in the instability of the local structure of Glut1,which interferes with the functioning of the glucose transporter and hinders the uptake of glucose by the extracellular environment.This study provides a more detailed explanation of the reason why the R126C mutation leads to the decreased structural stability of Glut and the-ability to transport substrates at the atomic level,which will help to further understand the structure of Glut and provide a theoretical basis for the prediction and treatment of the disease.