Molecular Dynamics Simulations Of Biological Macromolecules

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pH dependance of a Na channel Sodium ion channels play an important role in electrical signaling in cells; as such they are the targets of many drugs, as well as naturally occurring toxins from plant and animal sources. Inhibition and/or improper functioning of sodium channels due to mutation can lead to disease. In bacterial voltage gated sodium channels, the passage of sodium ions through the pore is controlled by a selectivity filter (SF) comprised of four glutamate residues. The number of ions bound in the channel can vary, but is about 2 on average. We have shown with MD simulations at constant pH and with free energy perturbation that the pKa values of the four SF glutamate residues depend on the number of ions bound in the channel. With 2 or 3 ions bound, at physiological pH, the SF is most likely in fully deprotonated state, and possibly also the singly protonated state. With 1 or 0 ions bound, the doubly protonated state can also get populated. Based on the MD simulations of the fully open channel, we have further shown that the conductance of the channel decreases with each proton bound to the SF. Thus the conductance of the channel is pH dependent, and decreases with lowering of pH. We also show that the conductance depends on the lipid composition of the membrane. The mechanisms involved in modulation of the channel conductances involves changes in the radius of the gate and the SF, as well as the electric field in the pore. Mechanism of degradation of Histatin 5 peptide by Secreted Aspartic Protease (SAPS) of C. Albicans This project started with testing the double link atom (DLM) methodology to treat the boundary between QM and MM regions on Amino Acids. The charge of MM link atom for Amino Acids was optimized by minimizing the difference between the dipole of the molecule before and after removing the QM part. Next, mechanism of cleavage of Hst5 by Sap-2 the major produced Aspartic Protease of C. Albicans and the effect of mutation on the cleavage process was studied with QM/MM methodology. Docking the peptide to the active site of Saps is performed by restraining the distance between the active site aspartic (Asp) residues and the lysine (Lys) residue on Hst5. To find difference between binding of Hst5 and its mutants to SAP we performed replica exchange umbrella sampling (REUS) of the peptide by pulling the peptide from active site to bulk water phase. The results showed a -11 kcal/mol free energy of binding for Hst5 to SAP. Initial conformation for quantum region is obtained by putting a harmonic restraint between Lys of the peptide and Asp of the enzyme and it is observed that water molecules occupy the active site. Intermediate and product states of the reaction are produced by restrained QMMM optimization with DFT level of theory for quantum region. The mechanism involves a tetrahedral gem-diol intermediate state which then leads to amide bond hydrolysis. A replica path method in CHARMM was used to find transition state and minimum energy path (MEP) of the reaction with Hartree Fock (HF) level of theory of QM region with 6-31G basis set. It was found that the formation of the intermediate state is the limiting step of the reaction. However, a higher level of theory and a more complex basis set is now being used to confirm these results. In the next step, we will use the optimized reaction path to start an off-path sampling which allows us to find the free energy of the reaction. Critical Sequence Hot-spots for Binding of nCOV-2019 to ACE2 as Evaluated by MD simulations: The novel coronavirus (nCOV-2019) outbreak has put the world on edge, causing millions of cases and hundreds of thousands of deaths all around the world, as of June 2020, let alone the societal and economic impacts of the crisis. The spike protein of nCOV-2019 resides on the virions surface mediating coronavirus entry into host cells by binding its receptor binding domain (RBD) to the host cell surface receptor protein, angiotensin converter enzyme (ACE2). In this study we have provided a detailed structural mechanism of how nCOV-2019 recognizes and establishes contacts with ACE2 and its difference with an earlier coronavirus SARS-COV in 2002 via extensive molecular dynamics (MD) simulations. Our results showed that nCOV-2019 RBD binds ACE2 with a significantly higher affinity () than SARS-COV which correlates with its higher infection rate. A per-residue free energy decomposition pinpointed the critical role of nCOV-2019 RBD residues Lys417, Tyr505, Gln498, Gln493 in binding ACE2. Numerous mutations have been identified in the RBD of nCOV-2019 strains isolated from humans in different parts of the world. In this study, we investigated the effect of these mutations as well as other Ala-scanning mutations on the stability of RBD/ACE2 complex. It is found that most of the naturally occurring mutations to the RBD either slightly strengthen or have the same binding affinity to ACE2 as the wild-type nCOV-2019. This means the virus had sufficient binding affinity to its receptor at the beginning of the crisis. This also have implications for any vaccine design endeavors since these mutations could act as antibody escape mutants. Furthermore, in-silico Ala-scanning and long-timescale MD simulations, highlight the crucial role of the residues at the interface of RBD and ACE2 that may be used as potential pharmacophores for any drug development endeavors. From an evolutional perspective, this study also identifies how the virus has evolved from its predecessor SARS-COV and how it could further evolve to become even more infectious. Exploring dynamics and network analysis of spike glycoprotein in SARS-COV-2 The ongoing pandemic caused by coronavirus SARS-COV-2 continues to rage with devastating consequences on human health and global economy. A spike glycoprotein on the surface of coronavirus mediates its entry into host cells and is the target of all antibody design efforts to neutralize the virus. The glycan shield of the spike helps the virus to evade the human immune response by providing a thick sugar-coated barrier against any antibody. To study the dynamic motion of glycans in the spike protein we performed microsecond-long MD simulation on the spike protein in two different states that correspond to the receptor binding domain in open or closed conformations. Analysis of this microsecond-long simulation revealed a scissoring motion on the N-terminal domain of neighboring monomers in the trimer. Role of multiple glycans in shielding of spike protein in different regions were uncovered by a network analysis. Centrality measurements in graph theory helped us identify glycans that play local or global roles in the network. It was found that the stalk region glycans have high local centralities which give rise to an effective shielding of this domain. On the other hand, breaches can be found in the apex of the spike protein for antibodies to bind and neutralize the virus. Role of several glycan such as N234 and N165 were pinpointed in the network of glycan. Microdomains of glycans were identified featuring a high degree of intra-communication in these microdomains and therefore most antibodies would bind to regions between these microdomains. An antibody overlap analysis revealed the glycans microdomains as well as individual glycans that inhibit access to the antibody epitopes on the spike protein. Our analysis showed that the spike protein is more vulnerable to antibodies when the RBD is in the open state. Overall, the results of this study provide detailed understanding of the spike glycan shield which must be considered for any rational antibody design project.