In vitro characterization of ionizing radiation resistance protein YejH

Abstract

DNA damage resulting from exposure to Ionizing Radiation (IR) is fatal unless it is repaired. Organisms have evolved mechanisms to maintain genomic integrity upon insult. In a screen for genes that contribute to IR resistance, gene of unknown function, yejH, was identified. Cells deleted for yejH are nearly a 100‐fold more sensitive to IR exposure than wild type. YejH is a predicted helicase; the primary amino acid sequence contains the seven conserved domains of Superfamily 2 helicases. To support our hypothesis that YejH is a DNA repair enzyme, both DNA binding and DNA‐dependent ATPase activity were assayed. We found that YejH is DNA‐independent ATPase, however it binds DNA in the presence of ATP and non‐hydrolyzable ATPγS. Using Tandem Affinity Purification, single‐stranded DNA binding protein (SSB) was identified as a binding partner of YejH. SSB binds single‐stranded DNA (ssDNA) and interacts with a growing list of proteins, thus localizing DNA metabolism proteins to replication forks and damaged DNA. This work aims to characterize YejH in vitro and determine how SSB affects YejH activity. As we understand more about the proteins that interact with YejH and how YejH activity is affected, we become closer to understanding the role of YejH in surviving IR exposure.