Insights Into The Mechanistic Origin Of Different Binding Affinities Of SARS-CoV And SARS-CoV-2 Spike RBDs To Human ACE2 And The Cold-Adaptation Mechanism Of Serine Hydroxymethyltransferase Using The Computational Methods

The dissertation contains two studies,which are:1.Mechanistic origin of different binding affinities of SARS-CoV and SARS-CoV-2 spike RBDs to human ACE2The receptor-binding domain(RBD)of the SARS-CoV-2 spike protein mediates viral entry into host cells through binding to the cell-surface receptor angiotensinconverting enzyme 2(ACE2).It has been shown that SARS-CoV-2 RBD(RBDCoV2)has a higher binding affinity to human ACE2 than its highly homologous SARS-CoV RBD(RBDcov),for which the mechanistic reasons still remain to be elucidated.Here,we used the multiple-replica molecular dynamics(MD)simulations,molecular mechanics Poisson-Boltzmann surface area(MM-PBSA)binding free energy calculations,and interface residue contact network(IRCN)analysis approach to explore the mechanistic origin of different ACE2 binding affinities of these two RBDs.The results demonstrate that,when compared to the RBDCoV2-ACE2 complex,the RBDCoV-ACE2 complex features the enhanced overall structural fluctuations and inter-protein positional movements and increased conformational entropy and diversity.The inter-protein electrostatic attractive interactions are a dominant force in determining the high ACE2 affinities of both RBDs,while the significantly strengthened electrostatic forces of attraction of ACE2 to RBDCoV2 determine the higher ACE2 binding affinity of RBDCoV2 than of RBDcov.Comprehensive comparative analyses of the residue binding free energy components and IRCNs reveal that,although any RBD residue substitution involved in the charge change can significantly impact the inter-protein electrostatic interaction strength,it is the residue changes at the RBD interface that lead to the overall stronger electrostatic attractive force of RBDCoV2 with ACE2,which not only tightens the interface packing and suppresses the dynamics of RBDCoV2-ACE2,but also enhances the ACE2binding affinity of RBDCoV2.Since the RBD residue substitutions involving the gain/loss of the positively/negatively charged residues,in particular those near or at the binding interfaces with the potential to form hydrogen bonds and/or salt bridges with ACE2,can greatly enhance the ACE2 binding affinity,the SARS-CoV-2 variants carrying such mutations should be paid special attention to.2.Exploring the cold-adaptation mechanism of serine hydroxymethyltransferase by comparative molecular dynamics simulationsCold-adapted enzymes feature a lower thermostability and higher catalytic activity compared to their warm-active homologues,which are considered as a consequence of increased flexibility of their molecular structures.The complexity of the(thermo)stability-flexibility-activity relationship makes it difficult to define the strategies and formulate a general theory for enzyme cold adaptation.Here,the psychrophilic serine hydroxymethyltransferase(pSHMT)from Psychromonas ingrahamii and its mesophilic counterpart,mSHMT from Escherichia coli,were subjected to μs-scale multiple-replica molecular dynamics(MD)simulations to explore the cold-adaptation mechanism of the dimeric SHMT.The comparative analyses of MD trajectories reveal that pSHMT exhibits larger structural fluctuations and inter-monomer positional movements,a higher global flexibility,and considerably enhanced local flexibility involving the surface loops and active sites.The largest-amplitude motion mode of pSHMT describes the trends of intermonomer dissociation and enlargement of the active-site cavity,whereas that of mSHMT characterizes the opposite trends.Based on the comparison of the calculated structural parameters and constructed free energy landscapes between the two enzymes,we discuss in-depth the physicochemical principles underlying the stability-flexibility-activity relationships and conclude that ⅰ)pSHMT adopts the global-flexibility mechanism to adapt to the cold environment and,ⅱ)optimizing the protein-solvent interactions and loosening the inter-monomer association are the main strategies for pSHMT to enhance its flexibility.