Theoretical Studies On The Effect Of Protein Secondary Structure Changes Induce Its Activity Using Molecular Dynamic Simulations

Protein is the basic substance for the living activities,and it participates in all aspects of life activities.The structure and function of proteins are determined by the tertiary structure of proteins consisting of secondary structures.Changes in secondary structure of proteins often affect the activity of proteins.Traditional experimental techniques of molecular biology can not detect the changes of secondary structure microscopically,nor can they analyze the molecular mechanism of secondary structure changes regulating protein activity.Molecular dynamic simulation is a new and cross-cutting biological method with the development of biotechnology and computer science.It has been applied to the study of catalytic mechanism of biological macromolecule,protein conformation change,enzyme-ligand interaction mechanism.It has played its unique advantages and has become an irreplaceable method in many aspects.In this study,several molecular simulation methods(molecular dynamics simulation,molecular docking,protein network analysis,binding free energy calculation)were used to explore several important proteins in depth,and the molecular mechanism of secondary structure of proteins regulating protein activity was discussed in depth.The specific research contents of this paper are as follows:1.Molecular mechanism of MEK1 secondary structure change affecting its catalytic activityApproximately 30% of all types of human cancers possess a constitutively activated the mitogen-activated protein kinase(MAPK)signaling pathway while MAP kinase 1(MEK1)is a critical component of this pathway.It has been reported mutations could improve the activity of MEK1 to result in cell proliferation and transformation,which is a known oncogenic event in various cancer types.In this study,eight molecular dynamics simulations,Molecular Mechanics Poisson-Boltzmann Surface Area(MM-PBSA),combined with protein structure network were performed to explore the mechanism that mutations activate MEK1.Protein structure networks and hydrogen bonds analysis demonstrated that active mutations broke the interaction between activation segments(Residues 216–222)and C-helix(Residues 105–121)in MEK1,leading to it transform inactive form to active form.Moreover,hydrogen bond analysis and MM-PBSA calculation indicated that activating mutations decrease the binding affinity between MEK1 and inhibitor to reduce the inhibitory effect of inhibitors.In addition,some active mutations cause structural changes in the Pro-rich loop(Residues 261–268)of MEK1.These changes may stabilize the interaction between the MEK1 mutants and the ligands by increasing the number of exposed hydrophobic residues in the active site of MEK1.Our results may provide useful theoretical evidences for the mechanism underlying the role of human MEK1 in human cancers.2.Molecular mechanism of zearalenone hydrolase secondary structure change affecting its degradation of ?-ZOLZearalenone hydrolase(ZHD)is the only reported ?/?-hydrolase that can detoxify zearalenone(ZEN).ZHD has demonstrated its potential as a treatment for ZEN contamination that will not result in damage to cereal crops.Recent researches have shown that the V153 H mutant ZHD increased the specific activity against ?-ZOL,but decreased its specific activity to ?-ZOL.To understand why V153 H mutation showed catalytic specificity for ?-ZOL,four molecular dynamics simulations combining with protein network analysis for wild type ZHD ?-ZOL,ZHD ?-ZOL,V153 H ?-ZOL,and V153 H ?-ZOL complexes were performed using Gromacs software.Our theoretical results indicated that the V153 H mutant could cause a conformational switch at the cap domain(Residues Gly161–Thr190)to affect the relative position catalytic residue(His242).Protein network analysis illustrated that the V153 H mutation enhanced the communication with the whole protein and residues with high betweenness in the four complexes,which were primarily assembled in the cap domain and residues Met241 to Tyr245 regions.In addition,the existence of ?-ZOL binding to V153 H mutation enlarged the distance from the OAE atom in ?-ZOL to the NE2 atom in His242,which prompted the side chain of His242 to the position with catalytic activity,thereby increasing the activity of V153 H on the ?-ZOL.Furthermore,?-ZOL could easily form a right attack angle and attack distance in the ZHD and ?-ZOL complex to guarantee catalytic reaction.The alanine scanning results indicated that modifications of the residues in the cap domain produced significant changes in the binding affinity for ?-ZOL and ?-ZOL.Our results may provide useful theoretical evidence for the mechanism underlying the catalytic specificity of ZHD.3.Molecular mechanisms of secondary structure change of Thermus thermophilus argonaute affecting catalytic activityThermus thermophiles Argonaute(Tt Ago)is a complex,which is consisted of 5?-phosphorylated guide DNA and a series of target DNA with catalytic activities at high temperatures.To understand why high temperatures are needed for the catalytic activities,three molecular dynamics simulations and binding free energy calculations at 310,324 and 338 K were performed for the Tt Ago-DNA complex to explore the conformational changes between 16-mer guide DNA/15-mer target DNA and Tt Ago at different temperatures.The simulation results indicate that a collapse of a small ?-strand(Residues Glu507-Arg513)at 310 K caused Glu512 to move away from the catalytic residues Asp546 and Asp478,resulting in a decrease in catalytic activity,which was not observed in the simulations at 324 and 338 K.The nucleic acid binding channel became enlarged at 324 and 338 K,thereby facilitating the DNA to slide in.Binding free energy calculations and hydrogen bond occupancy indicated that the interaction between Tt Ago and the DNA was more stable at 324 K and 338 K than at 310 K.The DNA binding pocket residues Lys575 and Asn590 became less solvent accessible at 324 and 338 K than at 310 K to influence hydrophilic interaction with DNA.Our simulation studies shed some light on the mechanism of Tt Ago and explained why a high temperature was needed by Tt Ago during gene editing.4.Exploration of catalytic selectivity for transaminase Btr R based on multiple molecular dynamics simulationsThe aminotransferase from Bacillus circulans(Btr R),which is involved in the biosynthesis of butirosin,catalyzes the pyridoxal phosphate(PLP)-dependent transamination reaction to convert valienone to ?-valienamine(a new ?-glycosidase inhibitor for the treatment of lysosomal storage diseases)with an optical purity enantiomeric excess value.To explore the stereoselective mechanism of valienamine generated by Btr R,multiple molecular dynamics simulations(MD)were performed for the Btr R/PLP/valienamine and Btr R/PLP/?-valienamine complexes.The theoretical results showed that ?-valienamine could make Btr R more stable and dense than valienamine.?-valienamine could increase the hydrogen bond probability and decrease the binding free energy between coenzyme PLP and Btr R by regulating the protein structure of Btr R,which was conducive to the catalytic reaction.?-valienamine maintained the formation of cation-p interactions between basic and aromatic amino acids in Btr R,thus enhancing its stability and catalytic activity.In addition,CAVER analysis revealed that ?-valienamine could make the tunnel of Btr R wider and straight,which was propitious to the removal of products from Btr R.Steered MD simulation results showed that valienamine interacted with more residues in the tunnel during dissociation compared with ?-valienamine,resulting in the need for a stronger force to be acquired from Btr R.Taken together,Btr R was more inclined to catalyze the substrates to form ?-valienamine,either from the point of view of catalytic reaction or product removal.