Using phage display to identify potential interacting regions of the varicella-zoster virus portal and scaffold proteins

Varicella-zoster virus (VZV) is an alphaherpesvirus that is the causative agent of varicella (chicken pox) and herpes zoster (shingles). Current treatments include nucleoside analogs such as acyclovir and its derivatives. These drugs inhibit viral replication by targeting the viral DNA polymerase. However, there are numerous additional, potential virus specific targets within the virus replication process. Recently, van Zeijl et al. and Visalli et al. described novel compounds that prevent the cleavage and packaging of viral genomic DNA into the provirus capsid (encapsidation) by targeting the capsid portal proteins. The studies performed in this thesis were designed to investigate potential interacting regions of the VZV portal (pORF54) and scaffold (pORF33.5) proteins. Little is known about the interaction of portal and scaffold proteins in VZV, however the interaction of these two proteins has been studied in a close relative, herpes simplex virus type 1 (HSV-1). A defined region of the HSV-1 scaffold protein (pORF26.5) that interacts with the portal protein (pUL6) was identified by Singer et al. These previous studies showed that a 12 amino acid stretch of the HSV-1 scaffold protein was essential for binding to the HSV-1 portal protein. This data was used to design a VZV scaffold peptide to use as a target in a procedure called “phage display panning”. Individual phage that showed affinity through a series of panning steps were amplified in E.coli and the DNA sequenced to determine the unique 12-mer amino acid stretch expressed by each individual phage isolate. Sequences were scored based on homology of the 12-mer peptides to pORF54. Several phage were isolated at random from the original, un-panned phage library and used as controls (control). Four phage from the panning experiments gave homology scores greater than the control phage. The region of homology for the four peptide sequences was examined in the context of the portal structure and location within the capsid. Additionally, phage sequences were compared to each other to identify trends in the amino acids sequences. This analysis was used to identify amino acids that might have increased affinity for one or more regions of pORF54. A core region of homology was identified within the phage sequences that included large polar amino acids. Many of the sequences contained three or more consecutive large polar amino acids histidine (H), arginine (K), and lysine (K)