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dc.contributor.authorWalshe, James Lachlan
dc.date.accessioned2019-06-28
dc.date.available2019-06-28
dc.date.issued2019-06-28
dc.identifier.urihttp://hdl.handle.net/2123/20650
dc.description.abstractRegulated transcription termination provides an efficient and responsive means to control gene expression. In bacteria, intrinsic termination occurs through the formation of an RNA stem-loop (T-loop), which disrupts the RNA polymerase elongation complex, resulting in its dissociation from the DNA template. Bacteria have a number of strategies for over-riding intrinsic T-loops, one of which is the formation of mutually exclusive RNA structures. Proteins that promote the formation of alternative RNA structures are known as anti-terminators. The ANTAR domains are a family of RNA binding anti-terminators that are most commonly associated with two components signalling (TCS) pathways. Unlike the canonical DNA binding output domains of TCS proteins, ANTAR domains facilitate their response by binding to RNA. EutV from E. faecalis is an ANTAR domain protein that controls gene expression of the eut operon. EutV is a model protein for studying ANTAR induced anti-termination. The ANTAR domain binding motif consists of dual RNA hexaloops, with the second hexaloop overlapping with the 5’ end of the intrinsic T-loop. Despite the well-defined binding motif, it is unknown at the molecular level how the ANTAR domains interact with RNA. In this work we have determined the 2.16 Å resolution crystal structure of the ANTAR domain protein EutV in its RNA-free form. The dimeric structure of EutV shows a high degree of similarity to the previously characterised AmiR, suggesting a common dimeric organisation for these domains. We went on to solve the structure of EutV bound to RNA, which represents the first reported structure of an ANTAR domain bound to its cognate RNA binding motif. Our studies highlight the key interactions between conserved EutV residues and the RNA, as well as protein conformational changes undergone upon RNA binding. These interactions were validated by alanine mutagenesis, which allowed us to propose an updated model for ANTAR domain anti-termination.en_AU
dc.publisherUniversity of Sydneyen_AU
dc.publisherFaculty of Scienceen_AU
dc.publisherSchool of Life and Environmental Sciencesen_AU
dc.rightsThe author retains copyright of this thesis. It may only be used for the purposes of research and study. It must not be used for any other purposes and may not be transmitted or shared with others without prior permission.en_AU
dc.subjectANTARen_AU
dc.subjectRNA bindingen_AU
dc.subjectanti-terminationen_AU
dc.subjectEutVen_AU
dc.titleStructural Characterisation of an ANTAR Domain Anti-Terminator Protein Bound to RNAen_AU
dc.typePhD Doctorateen_AU
dc.type.pubtypeDoctor of Philosophy Ph.D.en_AU
dc.description.disclaimerAccess is restricted to staff and students of the University of Sydney . UniKey credentials are required. Non university access may be obtained by visiting the University of Sydney Library.en_AU


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