Faculty Mentor(s)

Drs. Virginia McDonough Stukey and Joseph Stukey

Document Type

Poster

Event Date

4-11-2014

Abstract

A bacteriophage is a virus that infects and reproduces in bacteria. During productive infections—those that result in construction and release of infectious phage particles—key host cell metabolic processes are modified by the infecting phage and redirected toward making new phage particles. Protein-protein interactions are likely involved in this process. In this work, gene 80 of mycobacteriophage Vix, a gene cytotoxic to host strain Mycobacterium smegmatis, was studied. Our hypothesis was that an interaction between the Vix80 gene product and a host cell protein caused growth inhibition. Comparative analysis of the Vix80 protein sequence shows a conserved domain of unknown function 2786 (DUF2786) near the N-terminus. The Vix80 gene was dissected, and the N-terminal 66 residues, encompassing the entire DUF2786 domain, was found to be cytotoxic to M. smegmatis. DUF2786 was found to be homologous to a region of three M. smegmatis ORFs, two of which are related by alternate initiation points of the same sequence. Using in vitro protein pull-down and in vivo two-hybrid analyses, efforts are underway to look for possible interactions of Vix80 with itself and the three host proteins. As part of this process, different constructs of the Vix80 protein were expressed in Escherichia coli. We have determined that certain Vix80 constructs are also lethal to the E.coli host cell, while others permit growth, suggesting a conservation of cytotoxic function. Identifying the relevant phage and host gene products and understanding how phage exploit their host’s weaknesses could lead to new therapeutic options for many bacterial illnesses.

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