This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Abstract: We propose to perform voltage-driven non-equilibrium molecular dynamics (NEMD) simulations of a mutated alpha-hemolysin (a-HL) ion channel, using the molecular dynamics program NAMD (Kale et. al, 1999). The channel is first embedded in an artificial membrane, and solvated in 1M NaCl salt solution at neutral pH and equilibrated;then an external voltage potential is applied across the system. Additional constraints are placed on the system so that the residues at the protein-solvent interface are allowed to move freely while the rest of the protein remains frozen. These simulations are based on our previous calculations of the a-HL system, performed on the Lemieux Cluster at PSC (grant # mcb030021p), where we generated an I-V curve from -400mV to +400mV, in good correspondence with experimental results (Misakian and Kasianowicz, 2003). We collected concentration profiles as well as a 1-D potential of mean force (PMF) profile along the central pore axis for both chloride and sodium ions, which revealed a distinct preference for chloride ions in the narrow regions of the pore, including the most constricted section of the pore, which serves as a weak selectivity filter favoring anions. Additionally, the results revealed a site along the lining of the ion channel (at lys147) which might provide a good candidate for site-directed mutagenesis in order to change the permeant properties of the system from anion to cation selective. We would like to test this by generating I-V data at +200mV and -200 mV using mutated a-HL structures with one, three and seven lys147 ->ser point mutations. These calculations, consisting of equilibration and I-V calculations, would be most effectively carried out on the Bigben cluster at Pittsburgh Supercomputing Center, due to its scalable architecture and large number of available CPUs. Each of the I-V data points should require approximately 5000 hours of processor time on Bigben, for a total request of 30,000 service units. Kale, L., R. Skeel, M. Bhandarkar, R. Brunner, A. Gursoy, N. Krawetz, J. Phillips, A. Shinozaki, K. Varadarajan, and K. Schulten. 1999. NAMD2: Greater scalability for parallel molecular dynamics. Journal of Computational Physics 151: 283-312. Misakian M. and J. J. Kasianowicz. 2003. Electrostatic control of ion transport through the -hemolysin channel. Journal of Membrane Biology, 195 (3), 137-146.
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