Biochemical and genetic analysis of Lhr, a mysterious helicase/glycosylase conserved across species
Publication Date
July 31, 2023
Creator
Abstract
Changes to DNA structure frequently inhibit essential processes such as DNA replication. Overcoming replication blockage or collapse requires replication-coupled DNA repair enzymes that catalyse removal of aberrant DNA structures and chemically modified bases. The Lhr family of helicases are found throughout archaea, including in the Heimdall- and Nano- archaeota, and are present in several bacterial clades. This family can be divided into ‘Lhr-core’ and ‘Lhr-extended’ protein variants with the latter containing an as yet uncharacterised extended C-terminal domain.
Through genetic analysis we identified an expression phenotype of archaeal Lhr identical to the replication-coupled DNA repair enzymes Hel308 and RecQ and implicated bacterial Lhr in a novel mutation repair pathway and in overcoming oxidative stress through interaction with a Rad51 paralogue.
In vitro analysis demonstrated archaeal Lhr preferential targeting of replication fork structures through ATP-independent binding causing melting/distortion of the branch point. This allowed loading for directional translocation and unwinding through the ‘parental’ DNA strands. Characterisation of bacterial Lhr-CTD revealed a newly identified d-uracil DNA glycosylase activity, building an emerging story about the contribution of Lhr and its associated proteins in prokaryotic DNA repair. Further context is afforded through phylogenetic analysis of RecA/Rad51 family proteins revealing the emergence of protein sub-families.
Here we present a substantial breakthrough in the study of Lhr proteins, implicating them for direct involvement in replication-coupled repair and a wider role in base excision repair.
Item Type
ethesis
Thesis Type
PhD
Supervisors
Subjects (LC)
Associated Schools / Departments
School of Life Sciences
eprints ID
72474
UoN Repository URI
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Name
Ryan Buckley_ PhD Thesis_4342175.pdf
Type
Full-text
Description
Examined. PhD thesis corrected as discussed during viva/thesis defence.
Size
13.15 MB
Format
Adobe PDF
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