The crystal structure of bacteriophage λ RexA provides novel insights into the DNA binding properties of Rex-like phage exclusion proteins

Journal Article · 29 March 2024 · Nucleic Acids Research

DOI: https://doi.org/10.1093/nar/gkae212 · OSTI ID:2331370

Adams, Myfanwy C.; Schiltz, Carl J.; Sun, Jing; Hosford, Christopher J.; Johnson, Virginia M.; Pan, Hao; Borbat, Peter P.; Freed, Jack H.; Thomason, Lynn C.; Court, Carolyn; Court, Donald L.; Chappie, Joshua S.

Abstract RexA and RexB function as an exclusion system that prevents bacteriophage T4rII mutants from growing on Escherichia coli λ phage lysogens. Recent data established that RexA is a non-specific DNA binding protein that can act independently of RexB to bias the λ bistable switch toward the lytic state, preventing conversion back to lysogeny. The molecular interactions underlying these activities are unknown, owing in part to a dearth of structural information. Here, we present the 2.05-Å crystal structure of the λ RexA dimer, which reveals a two-domain architecture with unexpected structural homology to the recombination-associated protein RdgC. Modelling suggests that our structure adopts a closed conformation and would require significant domain rearrangements to facilitate DNA binding. Mutagenesis coupled with electromobility shift assays, limited proteolysis, and double electron–electron spin resonance spectroscopy support a DNA-dependent conformational change. In vivo phenotypes of RexA mutants suggest that DNA binding is not a strict requirement for phage exclusion but may directly contribute to modulation of the bistable switch. We further demonstrate that RexA homologs from other temperate phages also dimerize and bind DNA in vitro. Collectively, these findings advance our mechanistic understanding of Rex functions and provide new evolutionary insights into different aspects of phage biology.

• Sponsoring Organization: USDOE
• Grant/Contract Number: AC02-06CH11357
• OSTI ID: 2331370
• Journal Information: Nucleic Acids Research, Journal Name: Nucleic Acids Research Journal Issue: 8 Vol. 52; ISSN 0305-1048
• Publisher: Oxford University PressCopyright Statement
• Country of Publication: United Kingdom
• Language: English