RESUMEN
Binding of herpes simplex virus (HSV) glycoprotein D (gD) to a cell surface receptor is required to trigger membrane fusion during entry into host cells. Nectin-1 is a cell adhesion molecule and the main HSV receptor in neurons and epithelial cells. We report the structure of gD bound to nectin-1 determined by x-ray crystallography to 4.0 Å resolution. The structure reveals that the nectin-1 binding site on gD differs from the binding site of the HVEM receptor. A surface on the first Ig-domain of nectin-1, which mediates homophilic interactions of Ig-like cell adhesion molecules, buries an area composed by residues from both the gD N- and C-terminal extensions. Phenylalanine 129, at the tip of the loop connecting ß-strands F and G of nectin-1, protrudes into a groove on gD, which is otherwise occupied by C-terminal residues in the unliganded gD and by N-terminal residues in the gD/HVEM complex. Notably, mutation of Phe129 to alanine prevents nectin-1 binding to gD and HSV entry. Together these data are consistent with previous studies showing that gD disrupts the normal nectin-1 homophilic interactions. Furthermore, the structure of the complex supports a model in which gD-receptor binding triggers HSV entry through receptor-mediated displacement of the gD C-terminal region.
Asunto(s)
Moléculas de Adhesión Celular/química , Herpesvirus Humano 1/química , Receptores Virales/química , Proteínas del Envoltorio Viral/química , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Herpesvirus Humano 1/fisiología , Humanos , Nectinas , Unión Proteica , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Receptores Virales/genética , Receptores Virales/metabolismo , Relación Estructura-Actividad , Proteínas del Envoltorio Viral/genética , Proteínas del Envoltorio Viral/metabolismo , Internalización del VirusRESUMEN
Histone deacetylase 4 (HDAC4) is a histone deacetylase profoundly involved in cell differentiation and in the pathogenesis of cancer. The histone deacetylase inhibitors are a new, promising class of anticancer agents. The screening of molecular interactions involving determination of the affinity of drug candidates is an integral part of the drug discovery process. Here we report the development of an assay using surface plasmon resonance for the analysis of HDAC4-small molecule interactions. We describe a new cloning and purification strategy that can be used to set up surface plasmon resonance experiments with other recombinant proteins.