Expression, isolation, and characterization of the Varicella-zoster virus capsid portal protein, pORF54

Alexander J Howard, Purdue University

Abstract

The need for alternative drug therapies for herpesviral infections is evidenced by the generation of resistance to current drug compounds in several human herpesviruses, including HSV-1 and VZV. Previous research has indicated a novel class of thiourea replication inhibitors target the putative portal proteins of HSV-1 and Varicella-zoster. The VZV portal homolog, encoded by the ORF54 gene, was used to generate a GST-pORF54 fusion protein. Purified GST-pORF54 was used to generate a polyclonal guinea pig antiserum. The antiserum detected an 86.8 kDa polypeptide in infected cells that corresponds with the predicted size of the putative VZV portal protein, pORF54. In addition, a protein of the same size was observed in western blot analysis of sucrose density gradient purified VZV virions. Full length pORF54 was expressed in a recombinant baculovirus system. Sucrose density gradient fractionation resulted in samples containing enriched amounts of pORF54. Transmission electron microscopy indicated that these fractions contained structures that form homomultimeric VZV portal protein complexes. The results suggest the VZV ORF54 gene encodes a protein expressed in infected cells and assembles into higher order structures to form the VZV capsid portal protein. This is the first study to characterize the VZV portal protein via electron microscopy, the largest viral portal isolated to date.

Degree

M.S.

Advisors

Visalli, Purdue University.

Subject Area

Molecular biology|Medicine|Virology

Off-Campus Purdue Users:
To access this dissertation, please log in to our
proxy server
.

Share

COinS