title: Impact of calcium on N1 influenza neuraminidase dynamics and binding free energy.
abstract: The highly pathogenic influenza strains H5N1 and H1N1 are currently treated with inhibitors of the viral surface protein neuraminidase (N1). Crystal structures of N1 indicate a conserved, high affinity calcium binding site located near the active site. The specific role of this calcium in the enzyme mechanism is unknown, though it has been shown to be important for enzymatic activity and thermostability. We report molecular dynamics (MD) simulations of calcium-bound and calcium-free N1 complexes with the inhibitor oseltamivir (marketed as the drug Tamiflu), independently using both the AMBER FF99SB and GROMOS96 force fields, to give structural insight into calcium stabilization of key framework residues. Y347, which demonstrates similar sampling patterns in the simulations of both force fields, is implicated as an important N1 residue that can "clamp" the ligand into a favorable binding pose. Free energy perturbation and thermodynamic integration calculations, using two different force fields, support the importance of Y347 and indicate a +3 to +5 kcal/mol change in the binding free energy of oseltamivir in the absence of calcium. With the important role of structure-based drug design for neuraminidase inhibitors and the growing literature on emerging strains and subtypes, inclusion of this calcium for active site stability is particularly crucial for computational efforts such as homology modeling, virtual screening, and free energy methods.
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California, USA. mlawrenz@ucsd.edu
authors.forename: Morgan
authors.initials: M
authors.lastname: Lawrenz
authors.forename: Jeff
authors.initials: J
authors.lastname: Wereszczynski
authors.forename: Rommie
authors.initials: R
authors.lastname: Amaro
authors.forename: Ross
authors.initials: R
authors.lastname: Walker
authors.forename: Adrian
authors.initials: A
authors.lastname: Roitberg
authors.forename: J Andrew
authors.initials: JA
authors.lastname: McCammon
mesh_terms: D000998:Antiviral Agents / Q000737:chemistry; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D016000:Cluster Analysis; D006860:Hydrogen Bonding; D053118:Influenza A Virus, H1N1 Subtype / Q000201:enzymology*; D053124:Influenza A Virus, H5N1 Subtype / Q000201:enzymology*; D056004:Molecular Dynamics Simulation; D009439:Neuraminidase / Q000737:chemistry* / Q000378:metabolism; D053139:Oseltamivir / Q000737:chemistry; D011485:Protein Binding; D013816:Thermodynamics; D014764:Viral Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000998:Antiviral Agents; D014764:Viral Proteins; D053139:Oseltamivir; D009439:Neuraminidase; D002118:Calcium