Residue-based design of small molecule inhibitor for H1N1, H5N1 and H7N1 mutants.

Point mutations H274Y and N294S can lead to resistance of influenza virus strains to some drug molecules. Recently, a large number of experiments has focused on the many frameworks and catalytic residues thought to prevent the efficacy of anti-flu drugs. In the past, most research has considered the role of drugs in rigid proteins rather than in flexible proteins. In this study, we used molecular dynamics simulation (MD) combined with structure- and ligand-based drug design (SBDD and LBDD) methods to study dynamic interaction and protein dynamics correlation statistics between compounds and both the framework and catalytic residues in influenza virus N1 strains. Drug candidates were screened using the IC50 of the docking result predicted by support vector machine, multiple linear regression, and genetic function approximation (P < 0.001). As shown by MD, saussureamine C and diiodotyrosine have a protein dynamics correlation similar to that of sialic acid, and both can participate in hydrogen bond formation with loop, framework, and catalytic residues. Our in silico findings suggest that saussureamine C can inhibit H274Y and N294S mutants, and that diiodotyrosine can also inhibit N294S mutants. Therefore, the drugs saussureamine C and diiodotyrosine have the potential to produce inhibitory effects on wild-type influenza virus and some N1 mutants.

A screening assay for neuraminidase inhibitors using neuraminidases N1 and N3 from a baculovirus expression system.

CONTEXT: Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibitors. OBJECTIVE: Validate the use of recombinant neuraminidase expressed in baculovirus located on the viral surface capsule to develop a neuraminidase inhibitor screening assay. MATERIALS AND METHODS: Recombinant baculovirus particles displaying neuraminidase N1 and N3 were used as enzyme sources. The assay set-up required the use of 2'-(4-methylumbelliferyl)-α-D-acetyl neuraminic acid as substrate and oseltamivir carboxylate as benchmark inhibitor. RESULTS: The assay was set up in a standard 96-well plate. The within- and between-assay coefficients of variation were, on average, less than 10%. The 50% inhibitory concentration values of the inhibitor were in good agreement with those determined by independent kinetic experiments. DISCUSSION AND CONCLUSIONS: The assay showed satisfactory within- and between-assay repeatability. The obtained results suggest that recombinant baculovirus expressing neuraminidase located on the virus membrane capsule can be used to set up affordable and reliable neuraminidase inhibitors screening assays.