Testing the Behavior of a WYL Domain Protein as a Transcriptional Regulator for the BREX Phage Restriction System
Faculty Information
Brett Kaiser | Project-in-progress
Presentation Type
Individual
Presentation Format
Oral presentation
Start Date
10-5-2024 1:00 PM
End Date
10-5-2024 2:00 PM
Abstract or artist statement
Bacteriophage exclusion (“BREX”) is a relatively uncharacterized 6-gene phage restriction system found in a wide range of bacteria that performs two main activities: methylation of host DNA and phage restriction. It is currently unknown how BREX genes are regulated. We have identified a seventh gene that is associated with a subset of BREX systems, BrxR, that likely transcriptionally regulates the BREX system it is linked to. Biochemical and crystallographic analyses demonstrate that BrxR forms a homodimer and specifically binds a DNA target site upstream of its transcription start site using its wHTH domains. Additional examination of the protein's structure shows an abundance of polar groups in the WYL domain, which is located opposite the wHTH domain. We speculate that this region of BrxR functions as a sensor for phage infections by recognizing an unknown ligand produced during a phage infection, which could then induce a conformational change that alters DNA binding. We have attempted to determine the identity of the ligand by analyzing crystallographic data. Additionally, recent studies have shown that BrxR is not exclusive to BREX systems, but rather associates with various defense systems encoded within different bacterial genomes. We became interested in two highly related BrxR homologs that differ by only seven residues but bind with disparate affinities to their common DNA target. Here we test the contribution of the differing amino acids to DNA binding by systematically mutating these various positions. We hope that determining the significant amino acid positions that contribute to DNA binding will inform how BrxR DNA recognition occurs and how the BREX gene expression is regulated.
Keywords: Protein, Phage, Bacteria, Transcriptional Regulator, Defense System
Testing the Behavior of a WYL Domain Protein as a Transcriptional Regulator for the BREX Phage Restriction System
Bacteriophage exclusion (“BREX”) is a relatively uncharacterized 6-gene phage restriction system found in a wide range of bacteria that performs two main activities: methylation of host DNA and phage restriction. It is currently unknown how BREX genes are regulated. We have identified a seventh gene that is associated with a subset of BREX systems, BrxR, that likely transcriptionally regulates the BREX system it is linked to. Biochemical and crystallographic analyses demonstrate that BrxR forms a homodimer and specifically binds a DNA target site upstream of its transcription start site using its wHTH domains. Additional examination of the protein's structure shows an abundance of polar groups in the WYL domain, which is located opposite the wHTH domain. We speculate that this region of BrxR functions as a sensor for phage infections by recognizing an unknown ligand produced during a phage infection, which could then induce a conformational change that alters DNA binding. We have attempted to determine the identity of the ligand by analyzing crystallographic data. Additionally, recent studies have shown that BrxR is not exclusive to BREX systems, but rather associates with various defense systems encoded within different bacterial genomes. We became interested in two highly related BrxR homologs that differ by only seven residues but bind with disparate affinities to their common DNA target. Here we test the contribution of the differing amino acids to DNA binding by systematically mutating these various positions. We hope that determining the significant amino acid positions that contribute to DNA binding will inform how BrxR DNA recognition occurs and how the BREX gene expression is regulated.
Keywords: Protein, Phage, Bacteria, Transcriptional Regulator, Defense System