Molecular mechanisms of pilin glycosylation in Neisseria: a model system for protein glycosylation in bacteria. The disease causing bacteria Neisseria meningitidis and Neisseria gonorrhoeae are important human pathogens. Cell surface structures, called pili, are known to be important in allowing the bacteria to stick to host cells. Genetic and structural studies have identified that the protein subunits, which make up pili, are glycosylated - modified by the addition of sugars. Until recently ....Molecular mechanisms of pilin glycosylation in Neisseria: a model system for protein glycosylation in bacteria. The disease causing bacteria Neisseria meningitidis and Neisseria gonorrhoeae are important human pathogens. Cell surface structures, called pili, are known to be important in allowing the bacteria to stick to host cells. Genetic and structural studies have identified that the protein subunits, which make up pili, are glycosylated - modified by the addition of sugars. Until recently glycosylation of Gram-negative bacterial proteins was not thought to occur, however our recent work with these bacteria, and other groups studying Pseudomonas and Campylobacter, have shown that this process may be widespread. In our previous studies, we have identified and analysed a number of genes involved in pili glycosylation, in bacteria, which make known sugar structures. We have used this information to developed models for how the biochemistry and physiology of the glycosylation system may work. With a well-established structure and many genes already identified, glycosylation in Neisseria represents the best available model system to study this novel and important process. In the proposed study we describe experiments planned to test our models and reveal the molecular detail of this process. This study could lead to major advances in our understanding of this process and, when understood, may have future applications in biotechnology.Read moreRead less
Stuctural analysis of RNA polymerase elongation complexes. RNA polymerase (RNAP) is an essential enzyme in all living cells. Its role is to convert the genetic information stored in genes into a message that can be converted into protein. Many additional factors are required to ensure that this enzyme functions correctly in the cell. The aim of this project is to obtain structural information on a bacterial RNAP complexed with an essential transcription factor called NusA. Using this information ....Stuctural analysis of RNA polymerase elongation complexes. RNA polymerase (RNAP) is an essential enzyme in all living cells. Its role is to convert the genetic information stored in genes into a message that can be converted into protein. Many additional factors are required to ensure that this enzyme functions correctly in the cell. The aim of this project is to obtain structural information on a bacterial RNAP complexed with an essential transcription factor called NusA. Using this information, plus data already obtained on the structure of this enzyme complexed with another essential factor called sigma, we will design small molecules to inhibit the interaction of these essential factors with polymerase. These molecules will serve as leads for the development of new antibiotics.Read moreRead less
Host-pathogen interactions: the role of mimicry. The proposed research program, using a combination of structure and functional analysis will provide insight into the mechanism of nucleotide hydrolysis by the enzymes NTPDases. This study will not only improve our fundamental understanding of NTPDase action but could lead to the rational design of antimicrobials.
Investigation of a Novel Protein Implicated in Phosphate Metabolism in Bacteria. Phosphate is an important nutrient for all forms of life on Earth. A novel bacterial protein has been identified that appears to be important for the uptake or processing of phosphate, since mutants lacking the protein grow poorly inside certain cells of the human immune system (where phosphate levels are low) and in media containing low phosphate. The aims of this project are: to determine the role of the protein b ....Investigation of a Novel Protein Implicated in Phosphate Metabolism in Bacteria. Phosphate is an important nutrient for all forms of life on Earth. A novel bacterial protein has been identified that appears to be important for the uptake or processing of phosphate, since mutants lacking the protein grow poorly inside certain cells of the human immune system (where phosphate levels are low) and in media containing low phosphate. The aims of this project are: to determine the role of the protein by examining all phosphate containing molecules in our mutants; to determine its location in bacteria and functional domains; to identify other affected genes in our mutants; and, to find proteins that interact with this new protein. This project expects to demonstrate the importance of this protein in phosphate metabolism in bacteria.Read moreRead less
Novel mechanisms of bacterial arsenic metabolism - arsenate reduction and arsenite oxidation. Novel arsenic metabolising bacteria (i.e., arsenate respiring and arsenite oxidising), which are both phylogenetically and physiologically unique, have been isolated from arsenic-contaminated areas in Australia. The arsenate respiring bacterium, Chrysiogenes arsenatis, is of particular interest as it is the only organism reported able to respire with arsenate using the respiratory substrate acetate as t ....Novel mechanisms of bacterial arsenic metabolism - arsenate reduction and arsenite oxidation. Novel arsenic metabolising bacteria (i.e., arsenate respiring and arsenite oxidising), which are both phylogenetically and physiologically unique, have been isolated from arsenic-contaminated areas in Australia. The arsenate respiring bacterium, Chrysiogenes arsenatis, is of particular interest as it is the only organism reported able to respire with arsenate using the respiratory substrate acetate as the electron donor. It is proposed that physiological, biochemical and molecular biological studies be carried out to better understand the mechanisms by which these organisms metabolise arsenic. The knowledge gained from these studies will have worldwide application in the development of an arsenic bioremediation system.Read moreRead less
The protein O-glycosylation pathway in Neisseria meningitidis. Neisseria meningitidis causes bacterial meningitis, a sudden and severe disease of particular concern to children in both the developed and developing worlds. This project will contribute to an understanding of how these bacteria evade the immune system by modifying the proteins displayed on their surface, which will help in the development of a vaccine.
A Unique Target in the Purine Biosynthesis of the Pathogen Helicobacter pylori. The uptake systems of purine and analogues of the human pathogen Helicobacter pylori will be characterised because they can be utilised to introduce cytotoxic compounds into the cells. The first step in de novo purine biosynthesis of the bacterium is catalysed by two different enzymes, which are components of other biosynthetic pathways. These unique properties make them excellent potential therapeutic targets. Their ....A Unique Target in the Purine Biosynthesis of the Pathogen Helicobacter pylori. The uptake systems of purine and analogues of the human pathogen Helicobacter pylori will be characterised because they can be utilised to introduce cytotoxic compounds into the cells. The first step in de novo purine biosynthesis of the bacterium is catalysed by two different enzymes, which are components of other biosynthetic pathways. These unique properties make them excellent potential therapeutic targets. Their individual combined activities in purine biosynthesis will be characterised in situ and in vitro. Isogenic mutants with inactivated genes encoding for these enzymes will be constructed to investigate their role in the survival of the organism.Read moreRead less
Sensing atmosphere: Understanding the HNOX-protein gas-sensing capability and how it is affected by heme-oxidation. The project investigates how gas sensing heme-proteins from the novel HNOX (Heme-Nitric Oxide) family are able to discriminate between different gaseous ligands such as O2 and NO and how oxidation of the heme alters this response. The gas-sensing capability of the HNOX proteins is crucial for organisms ranging from bacteria to humans. Thus, understanding of these signalling mechani ....Sensing atmosphere: Understanding the HNOX-protein gas-sensing capability and how it is affected by heme-oxidation. The project investigates how gas sensing heme-proteins from the novel HNOX (Heme-Nitric Oxide) family are able to discriminate between different gaseous ligands such as O2 and NO and how oxidation of the heme alters this response. The gas-sensing capability of the HNOX proteins is crucial for organisms ranging from bacteria to humans. Thus, understanding of these signalling mechanisms will have a strong impact on many scientific fields from the control of pathogen growth to human blood pressure regulation. This collaboration will establish Australian scientists and as world-leading in the field of NO and redox signalling. This development will also be of substantial benefit for the training of the next generation of Australian students and scientists.Read moreRead less
Dissociation of a Tetrameric Enzyme with Interface-Targeted Peptides. With antibiotic resistance on the rise, there is an urgent need to develop new antibiotics and an equally urgent need to characterise new antibiotic targets. One such target is dihydrodipicolinate synthase (DHDPS) which catalyses the critical step in lysine and cell wall biosynthesis in bacteria. This proposal aims to generate new drugs targeting DHDPS for effective and rapid treatment of bacterial infections, including gastro ....Dissociation of a Tetrameric Enzyme with Interface-Targeted Peptides. With antibiotic resistance on the rise, there is an urgent need to develop new antibiotics and an equally urgent need to characterise new antibiotic targets. One such target is dihydrodipicolinate synthase (DHDPS) which catalyses the critical step in lysine and cell wall biosynthesis in bacteria. This proposal aims to generate new drugs targeting DHDPS for effective and rapid treatment of bacterial infections, including gastroenteritis. Recent statistics show that over 5 million Australians suffer from gastroenteritis each year and hospitalisation for this infection is nearly seven times higher for indigenous than non-indigenous children. Accordingly, this research has the potential to assure a healthier future for millions of Australians.Read moreRead less
How Bacteria Fold Virulence Factors to Cause Disease. Bacteria use folding enzymes to assemble proteins essential for cell integrity and pathogenicity. These foldases include the Disulphide bridge proteins, which catalyse the introduction of disulfide bonds. This project will study two important human pathogens, Salmonella Typhimurium and uropathogenic Escherichia coli, to address the fundamental and poorly understood questions of diversity of Dsb networks across bacterial pathogens and the role ....How Bacteria Fold Virulence Factors to Cause Disease. Bacteria use folding enzymes to assemble proteins essential for cell integrity and pathogenicity. These foldases include the Disulphide bridge proteins, which catalyse the introduction of disulfide bonds. This project will study two important human pathogens, Salmonella Typhimurium and uropathogenic Escherichia coli, to address the fundamental and poorly understood questions of diversity of Dsb networks across bacterial pathogens and the role of these foldases in virulence. The research will reveal how bacterial virulence factors are folded, identify novel targets for therapeutic intervention and provide the basis for structure-based design on new antimicrobials in the future. Read moreRead less