Abstract
To facilitate high-throughput biochemical analyses of membrane proteins, we have developed a novel display technology in a microarray format. Both single-pass (cluster of differentiation 4, CD4) and multiple-pass (G protein-coupled receptor 77, GPR77) human transmembrane proteins were engineered to be displayed in the membrane envelop of herpes simplex virions. These viruses produce large spherical virions displaying multiple copies of envelop proteins. Our aim was to engineer this virus to express these human proteins during the virus productive cycle and incorporate the human proteins into the virion during the assembly process. Another strategy presented includes engineering a fusion of glycoprotein C (gC), a major constituent of herpes simplex virus type 1 (HSV-1) virions, by hijacking the cis-acting signals to direct incorporation of the chimeric protein into the virion. The expression of the human proteins in infected cells, at the cell surface and in purified virions, is in the correct transmembrane orientation, and the proteins are biochemically functional. Purified virions printed on glass slides form a high-density Virion Display (VirD) Array, and the displayed proteins were demonstrated to retain their native conformations and interactions on the VirD Array judging by similar assays, such as antibody staining, as well as lectin and ligand binding. This method can be readily scaled or tailored for different modalities including a high-content, high-throughput platform for screening ligands and drugs of human membrane proteins.
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