PROJECT TITLE DESCRIPTION CHIEF INVESTIGATOR DETAILS

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1 PROJECT TITLE DESCRIPTION CHIEF INVESTIGATOR DETAILS Characterisation of the function of the essential transcription elongation factor NusA NusA is a highly conserved transcription elongation factor from bacteria that is essential for cell viability. We have developed a system for deleting wild-type NusA and simultaneously expressing mutant forms which will be used to determine which regions/functions of NusA are required for cell viability. This project will involve using this system to screen a panel of mutant NusA constructs. The results will help in the design of new approaches to target essential bacterial processes for the development of new antibiotics Understanding the role of the delta subunit of RNA polymerase RNA polymerase in bacteria contains multiple subunits. In Gram positive bacteria, in addition to α 2, β, β ω, there are additional δ and ε subunits. δ appears to be important in pathogenicity, although it remains unclear how it controls RNA polymerase activity. This project will build on previous work to identify the binding site of δ that will help us understand how it functions. The project will involve PCR, cloning, protein purification and protein binding assays. Making an antibiotic sensitive bacterium Antibiotic resistance is a growing global problem, and there are now bacterial strains resistant to all clinically useful drugs. In Gram negative bacteria the efflux protein TolC plays an important role in removing antibiotics from the cell, so making them resistant. In order to better understand how we can develop new drugs we will construct a deletion in the gene encoding TolC and compare its antibiotic sensitivity against the wild-type using established drugs as well as compounds currently under development in the lab Identification of a new transcription factor Transcription factors are needed to control the activity of RNA polymerase and in many bacteria have important roles in bacterial pathogenicity. The recently described factor CarD in the causative agent of tuberculosis is essential and plays a role in controlling the initiation of transcription. We have identified a gene in pathogenic gram positive bacteria similar to CarD and wish to establish if it is an ortholog. We will tag the candidate gene with green fluorescent protein (GFP) and establish whether the protein co-localises with RNA polymerase using live cell fluorescence microscopy.

2 Characterisation of the FtsK DNA translocase FtsK is a protein that links the key processes of bacterial cell division, chromosome segregation and chromosome unlinking. It acts as a molecular pump that moves DNA (whole chromosomes) away from the site of cell division and is involved in the untangling of the chromosomes and coordinates this with cell division. This project will further characterize the mechanism of this protein. Xer recombination in pathogenic bacteria DNA repair in bacteria with circular chromosomes can lead to a potentially lethal product, a chromosome dimer. This is resolved by a site-specific recombination event catalysed by the XerCD proteins. This project will characterise the XerCD genes from pathogenic bacteria: Staphylococcus aureus, Acinetobacter baumannii and Pseudomonas aeruginosa Activation of recombination: the XerD-FtsK interaction Xer recombination resolves chromosome dimers in bacterial cells. These dimers are lethal if unresolved. XerD requires interaction with the C-terminus of FtsK for catalytic activity, and this project will investigate this interaction. Biochemical and crystallographic studies will be combined to understand how activation occurs. The interaction between bowel-inflammation, hypoxia and bacterial pathogenesis The overall objective of this project is to examine whether bacterial universal stress response genes (usps) are the key switch that enables a benign, commensal bacterium to become a pathogen in the gastro-intestinal tract, in response to hypoxia and inflammation as seen in irritable bowel disease Identifying compounds to target sperm DNA damage Some toxic DNA damage in sperm is mediated by an enzyme, cyp2e1. We have an assay to test the efficacy of compounds in inhibiting the activity of the enzyme. This summer scholarship would allow the student to test compounds in the assay and then perform some DNA damage analysis. Dr Shaun Roman Shaun.Roman@newcastle.edu.auPhone: (02)

3 Regulation of cyp2e1 expression. The detoxifying enzyme, cyp2e1, demonstrates increased expression after cells are exposed to a toxicant. The summer scholarship would allow the student to use molecular biology tools to assess the expression of cyp2e1 and potential modifiers of expression (mirnas) in cells exposed to a variety of chemicals metabolised by cyp2e1 Dr Shaun Roman Shaun.Roman@newcastle.edu.au Phone: (02) Development of a metal ion biosensor in Acinetobacter This project combines synthetic biology with important basic research into an emerging human pathogen. It aims to develop metal ion biosensors that put fluorescent protein expression under the control of a metal sensing regulatory switch. The sensors will be used to identify factors involved in metal ion homeostasis in Acinetobacter. Due to the importance of metals for bacterial survival, these factors could be novel targets for the development of future antimicrobials. Developing a model to study the social behaviours of bacteria Many important bacterial activities, such as the colonisation of a host, require the coordinated production of secreted compounds by a community of cells. These secreted compounds are public goods because they benefit the whole community, but their production is energy intensive, and non-producing cheater cells can outcompete their neighbours. This project will develop a fluorescent reporter construct to identify cheater cells within a population and allow studies of social behaviours in bacteria. Examining the energy coupling mechanism of a novel multidrug efflux pump Multidrug efflux pumps sit in the bacterial membrane and catalyse the movement of antimicrobials out of the cell. The expression of just one of these proteins can facilitate cross resistance to several drug classes. My research group recently discovered a completely new family of multidrug efflux pumps, encoded by some of the deadliest and most drug resistant hospital pathogens. This project will investigate the energy coupling mechanism in a prototypical member of this family.

4 Synthesis and Evaluation of an Adsorbent Selective to 5-Fluorouracil and Derivatives Polymerisable 2,6-bis-(acrylamido) pyridine (BAAPy) can strongly associate in a 1:1 stoichiometric ratio with imide-containing templates, such as uracil and derivatives (e.g. 5-fluorouracil), by forming an array of hydrogen bonding interactions, i.e. donor-acceptor-donor/acceptor-donoracceptor (DAD/ADA) H-bonding. Selective polymeric adsorbents can be conveniently prepared by the process of molecular imprinting. Molecularly imprinted polymers or MIPs are robust, porous polymeric (i.e. plastic) molecular moulds with recognition capabilities specific for its target molecule. This project will involve the synthesis of BAAPy-based MIP adsorbent for 5-FU and evaluating its binding efficiency. Non-imprinted BAAPy-based adsorbent would also be capable of binding 5-FU and other uracil derivatives but MIP offers a more selective alternative which quite possibly will be able to separate various uracil derivatives. The MIP adsorbent will then be applied to the extraction of 5-FU from blood samples. Dr Clovia Holdsworth and A/Prof Jenny Schneider clovia.holdsworth@newcastle.edu.au Phone: (02) Towards the development of a MIP-Based Biosensor The project will involve the the synthesis of MIPs selective to cyanobacteria compounds, e.g. microcystein-lr and for potential use as a recognition element for an optical biosensor. The MIP will be attached to an optical fiber the signal transducer, thus, MIP will be prepared using controlled radical polymerisation. Dr Clovia Holdsworth and Prof Brett Neilan clovia.holdsworth@newcastle.edu.au Phone: (02) Synthetic developments towards new classes of BNCT agents This project aims to ultimately produce selective agents for cancer treatment to be used in boron neutron capture therapy, an emerging cancer management technique. Recent synthetic discoveries in the Simone group are allowing synthesis of new BNCT agents. Dr Michela Simone. michela.simone@newcastle.edu.au. Phone:(02) Expression of lanthipeptides This research project is focused on discovery and production of biologically active peptides known as lanthipeptides. Heterologous production of lanthipeptides in E. coli can overcome limitations present in the native organism, including low yield. A putative lanthipeptide gene cluster, identified in the cyanobacteria Nostoc punctiforme, has been isolated from a fosimd library. This summer scholarship will focus on the optimization of the isolated clone and expression of the product, allowing the student to engage with a developed project research whilst focusing on core molecular and microbiology skills. Professor Brett Neilan Brett.Neilan@newcastle.edu.auPhone: (02)

5 Identifying new pharmacological targets for prescription opioid dependence This project leads on from prelimary work supported by the PRC in establishing an animal model of opioid dependence in rats. Our pilot data indicates that a small but significant percentage of animal develop addiction like-traits to oxycodone - a common pain medication. Once established this model will be used to identify molecular targets for pharmacological interventions with a focus on the AKT-mTOR signalling. Assoc Prof Chris Dayas Christopher.Dayas@newcastle.edu.au and Prof Jenny Martin jen.martin@newcastle.edu.au The magnitude of the association between antipsychotic medication use in psychiatric admissions and the increased risk of sudden cardiac death. Rviewing medical records for data and inputting this data into an excel spreadsheet. Professor Jennifer Martin Dr Ingrid Berling Ingrid.Berling@hnehealth.nsw.gov.au Drug from dirt Finding new antibiotics is urgently needed because of antibiotic resistance in bacteria. Soil may help us to solve this problem. Many of the most widely used antibiotics have been derived from soil. For example, Penicillin is derived from Penicillium, a fungus/actinomycetes found in soil, and vancomycin is derived from a bacterium found in dirt. This project aims to prepare a review on the history of antibiotics derived from soil, and the current approaches to finding new antibiotics from soil. Professor Jennifer Martin, Professor Nanthi Bolan, nanthi.bolan@newcastle.edu.au Cannabis contamination Cannabis products can be contaminated with pesticides, heavy metals, plant growth hormones, and microbiological agents including mould and fungi. Contamination of cannabis occurs at various stages that include cultivation, processing and retailing. These contaminants are likely to impact the medicinal value of cannabis products. The project aims to undertake a review on the sources, distribution, bioavailability, toxicity, and risk assessment of various contaminants in cannabis products. Professor Jennifer Martin, Professor Nanthi Bolan, nanthi.bolan@newcastle.edu.au