ORCID Profile
0000-0002-0791-6850
Current Organisation
University of Adelaide
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Biochemistry and cell biology | Proteins and peptides | Synthetic biology | Analytical biochemistry |
Publisher: Elsevier BV
Date: 12-2009
DOI: 10.1016/J.PHARMTHERA.2009.09.004
Abstract: The aryl hydrocarbon receptor is a signal regulated transcription factor that has best been characterised as regulating the xenobiotic response to a variety of planar aromatic hydrocarbons. There is compelling evidence that it mediates most, if not all, of the toxic effects of dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin). Dioxin exposure results in a wide variety of toxic outcomes including severe wasting syndrome, chloracne, thymic involution, severe immune suppression, reduced fertility, hepatotoxicity, teratogenicity, tumour promotion and death. The pleiotropy of toxic outcomes implies the disruption of a wide range of normal physiological functions. The aryl hydrocarbon receptor has developmentally restricted expression as well as developmental defects in gene-targeted mice. It has a wide range of target genes that do not fit into the classical xenobiotic metabolising gene battery and has recently been shown to interact with NF-kappa B and the estrogen receptor. There is also evidence for its activation in the absence of exogenous ligand, all of which point to various roles outside xenobiotic metabolism. Ligands so far identified display differential activation potential with respect to receptor activity. This article addresses activities of the aryl hydrocarbon receptor that are outside the xenobiotic response. Known physiological roles are discussed as well as how their disruption contributes to the pleiotropic toxicity of TCDD.
Publisher: Elsevier BV
Date: 04-2009
Publisher: Oxford University Press (OUP)
Date: 03-08-2015
DOI: 10.1093/NAR/GKV778
Publisher: Portland Press Ltd.
Date: 05-10-2022
DOI: 10.1042/BST20220194
Abstract: Bacteria sense, interact with, and modify their environmental niche by deploying a molecular ensemble at the cell surface. The changeability of this exposed interface, combined with extreme changes in the functional repertoire associated with lifestyle switches from planktonic to adherent and biofilm states necessitate dynamic variability. Dynamic surface changes include chemical modifications to the cell wall export of erse extracellular biofilm components and modulation of expression of cell surface proteins for adhesion, co-aggregation and virulence. Local enrichment for highly repetitive proteins with high tandem repeat identity has been an enigmatic phenomenon observed in erse bacterial species. Preliminary observations over decades of research suggested these repeat regions were hypervariable, as highly related strains appeared to express homologues with erse molecular mass. Long-read sequencing data have been interrogated to reveal variation in repeat number in combination with structural, biophysical and molecular dynamics approaches, the Periscope Protein class has been defined for cell surface attached proteins that dynamically expand and contract tandem repeat tracts at the population level. Here, I review the erse high-stability protein folds and coherent interdomain linkages culminating in the formation of highly anisotropic linear repeat arrays, so-called rod-like protein ‘stalks’, supporting roles in bacterial adhesion, biofilm formation, cell surface spatial competition, and immune system modulation. An understanding of the functional impacts of dynamic changes in repeat arrays and broader characterisation of the unusual protein folds underpinning this variability will help with the design of immunisation strategies, and contribute to synthetic biology approaches including protein engineering and microbial consortia construction.
Publisher: Portland Press Ltd.
Date: 10-2015
DOI: 10.1042/BST20150088
Abstract: Staphylococcus aureus and Staphylococcus epidermidis are an important cause of medical device-related infections that are difficult to treat with antibiotics. Biofilms, in which bacteria are embedded in a bacterially-produced exopolymeric matrix, form on the surface of the implanted medical device. Our understanding of the molecular mechanisms underlying the initial surface attachment and subsequent intercellular interactions as the biofilm matures is improving. Biofilm accumulation can be mediated by a partially deacetylated form of poly-N-acetylglucosamine (PNAG) but, more recently, the role of bacterial surface proteins is being recognized. Here we describe the structure and function of two S. aureus cell surface proteins, FnBPA and SasG, implicated in host interactions and biofilm accumulation. These multifunctional proteins employ intrinsic disorder for distinct molecular outcomes. In the case of FnBPA, disorder generates adhesive arrays that bind fibronectin (Fn) in the case of SasG, disorder is, counterintuitively, used to maintain a strong extended fold.
Publisher: Proceedings of the National Academy of Sciences
Date: 03-10-2016
Abstract: Understanding the role played by disorder in biology is becoming increasingly important. Disordered proteins are central to signaling, development, initiation of transcription, and other vital cellular processes. How and why disordered proteins are used is not entirely clear, but disorder can be important in allostery, facilitate regulatory posttranslational modification, and allow rapid and specific but promiscuous binding. Here, our investigations of biofilm-promoting protein SasG illustrate that disorder can play another role. We show that the intrinsic disorder of one-half of the domains is important for imparting long-range cooperativity in folding of a large multidomain protein—allowing formation of a small local element of structure to precipitate cooperative folding of adjacent disordered domains across a length scale of ∼10 nm.
Publisher: Portland Press Ltd.
Date: 10-2015
DOI: 10.1042/BST20150088
Abstract: Staphylococcus aureus and Staphylococcus epidermidis are an important cause of medical device-related infections that are difficult to treat with antibiotics. Biofilms, in which bacteria are embedded in a bacterially-produced exopolymeric matrix, form on the surface of the implanted medical device. Our understanding of the molecular mechanisms underlying the initial surface attachment and subsequent intercellular interactions as the biofilm matures is improving. Biofilm accumulation can be mediated by a partially deacetylated form of poly-N-acetylglucosamine (PNAG) but, more recently, the role of bacterial surface proteins is being recognized. Here we describe the structure and function of two S. aureus cell surface proteins, FnBPA and SasG, implicated in host interactions and biofilm accumulation. These multifunctional proteins employ intrinsic disorder for distinct molecular outcomes. In the case of FnBPA, disorder generates adhesive arrays that bind fibronectin (Fn) in the case of SasG, disorder is, counterintuitively, used to maintain a strong extended fold.
Publisher: Springer Science and Business Media LLC
Date: 06-2015
DOI: 10.1038/NCOMMS8271
Abstract: Bacteria exploit surface proteins to adhere to other bacteria, surfaces and host cells. Such proteins need to project away from the bacterial surface and resist significant mechanical forces. SasG is a protein that forms extended fibrils on the surface of Staphylococcus aureus and promotes host adherence and biofilm formation. Here we show that although monomeric and lacking covalent cross-links, SasG maintains a highly extended conformation in solution. This extension is mediated through obligate folding cooperativity of the intrinsically disordered E domains that couple non-adjacent G5 domains thermodynamically, forming interfaces that are more stable than the domains themselves. Thus, counterintuitively, the elongation of the protein appears to be dependent on the inherent instability of its domains. The remarkable mechanical strength of SasG arises from tandemly arrayed ‘cl ’ motifs within the folded domains. Our findings reveal an elegant minimal solution for the assembly of monomeric mechano-resistant tethers of variable length.
Publisher: International Union of Crystallography (IUCr)
Date: 22-02-2012
Publisher: Cold Spring Harbor Laboratory
Date: 24-12-2020
DOI: 10.1101/2020.12.24.424174
Abstract: Changes at the cell surface enable bacteria to survive in dynamic environments, such as erse niches of the human host. Here, we reveal “Periscope Proteins” as a widespread mechanism of bacterial surface alteration mediated through protein length variation. Tandem arrays of highly similar folded domains can form an elongated rod-like structure thus variation in the number of domains determines how far an N-terminal host ligand binding domain projects from the cell surface. Supported by newly-available long-read genome sequencing data, we propose this new class could contain over 50 distinct proteins, including those implicated in host colonisation and biofilm formation by human pathogens. In large multi-domain proteins, sequence ergence between adjacent domains appears to reduce inter-domain misfolding. Periscope Proteins break this “rule”, suggesting their length variability plays an important role in regulating bacterial interactions with host surfaces, other bacteria and the immune system.
Publisher: International Union of Crystallography (IUCr)
Date: 30-06-2015
DOI: 10.1107/S1399004715009220
Abstract: The reduction of uridine to dihydrouridine at specific positions in tRNA is catalysed by dihydrouridine synthase (Dus) enzymes. Increased expression of human dihydrouridine synthase 2 (hDus2) has been linked to pulmonary carcinogenesis, while its knockdown decreased cancer cell line viability, suggesting that it may serve as a valuable target for therapeutic intervention. Here, the X-ray crystal structure of a construct of hDus2 encompassing the catalytic and tRNA-recognition domains (residues 1–340) determined at 1.9 Å resolution is presented. It is shown that the structure can be determined automatically by phenix.mr_rosetta starting from a bacterial Dus enzyme with only 18% sequence identity and a significantly ergent structure. The overall fold of the human Dus2 is similar to that of bacterial enzymes, but has a larger recognition domain and a unique three-stranded antiparallel β-sheet insertion into the catalytic domain that packs next to the recognition domain, contributing to domain–domain interactions. The structure may inform the development of novel therapeutic approaches in the fight against lung cancer.
Publisher: Proceedings of the National Academy of Sciences
Date: 22-04-2015
Abstract: RNA-binding proteins use erse mechanisms for generating specificity toward distinct RNA molecules. Different subfamilies of bacterial dihydrouridine synthases (Dus) modify specific uridines in tRNA, but the mechanism for selection of the target nucleotide is unknown. We solved crystal structures of the U16-specific Dus from Escherichia coli complexed with two different tRNAs. These structures reveal that the tRNA is bound in a completely different orientation from that observed in a U20-specific enzyme. The major reorientation of the substrate tRNA, driven by unique amino acid “binding signatures” and plasticity in the position of the C-terminal recognition domain, appears to be an evolutionary innovation to the known strategies that define specificity of enzymes toward tRNA.
Publisher: American Chemical Society (ACS)
Date: 17-06-2020
Publisher: American Chemical Society (ACS)
Date: 08-05-2017
Publisher: Elsevier BV
Date: 03-2023
Publisher: Proceedings of the National Academy of Sciences
Date: 09-12-2019
Abstract: Streptococcus groups A and B cause serious infections, including early onset sepsis and meningitis in newborns. Rib domain-containing surface proteins are found associated with invasive strains and elicit protective immunity in animal models. Yet, despite their apparent importance in infection, the structure of the Rib domain was previously unknown. Structures of single Rib domains of differing length reveal a rare case of domain atrophy through deletion of 2 core antiparallel strands, resulting in the loss of an entire sheet of the β-sandwich from an immunoglobulin-like fold. Previously, observed variation in the number of Rib domains within these bacterial cell wall-attached proteins has been suggested as a mechanism of immune evasion. Here, the structure of tandem domains, combined with molecular dynamics simulations and small angle X-ray scattering, suggests that variability in Rib domain number would result in differential projection of an N-terminal host-colonization domain from the bacterial surface. The identification of 2 further structures where the typical B-D-E immunoglobulin β-sheet is replaced with an α-helix further confirms the extensive structural malleability of the Rib domain.
Publisher: Oxford University Press (OUP)
Date: 08-11-2011
DOI: 10.1093/NAR/GKR906
Publisher: American Society for Pharmacology & Experimental Therapeutics (ASPET)
Date: 15-03-2004
Abstract: The aryl hydrocarbon receptor (AhR) is traditionally defined as a transcription factor activated by exogenous polyaromatic and halogenated aromatic hydrocarbon (PAH/HAH) ligands. Active AhR induces genes involved in xenobiotic metabolism, including cytochrome P4501A1, which function to metabolize activating ligands. However, recent studies implicate AhR in biological events that are apparently unrelated to the xenobiotic response, implying that endogenous activation mechanisms exist. Three AhR genes in zebrafish (Danio rerio) encode proteins that demonstrate differential activation in response to PAH/HAHs, with the nonresponsive drAhR1a having some sequence ergence from the PAH/HAH-responsive AhRs in the ligand binding domain (LBD). We used these differences to guide the mutagenesis of mouse AhR (mAhR), aiming to generate variants that functionally discriminate between activation mechanisms. We found substitution of histidine 285 in the LBD with tyrosine gave a receptor that could be activated by isopropyl-2-(1,3-dithietane-2-ylidene)-2-[N-(4-methylthiazol-2-yl)carbamoyl]acetate (YH439), a potential AhR ligand chemically distinct from classic PAH/HAH-type ligands, but prevented activation by both exogenous PAH/HAH ligands and the endogenous activation mimics of suspension culture and application of shear-stressed serum. The differential response of H285Y mAhR to YH439 suggests that this activator has a novel mode of interaction that tolerates tyrosine at position 285 in the LBD and is distinct from the binding mode of the well characterized PAH/HAH ligands. In support of this, the PAH-type antagonist 3',4'-dimethoxyflavone blocked mAhR activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin but not YH439. Furthermore, the strict correlation between response to exogenous PAH/HAH ligands and mimics of endogenous activation suggests that a PAH-type ligand may underpin endogenous mechanisms of activation.
Publisher: Cold Spring Harbor Laboratory
Date: 25-05-2021
DOI: 10.1101/2021.05.25.445580
Abstract: Biological engineers seek to have better control and a more complete understanding of the process of translation initiation within cells so that they may produce proteins more efficiently, as well as to create orthogonal translation systems. Previously, initiator tRNA variants have been created that initiate translation from non-AUG start codons, but their orthogonality has never been measured and the detailed characteristics of proteins produced from them have not been well defined. In this study we created an initiator tRNA mutant with anticodon altered to AAC to be complementary to GUU start codons. We deploy this i-tRNA(AAC) into E. coli cells and measure translation initiation efficiency against all possible start codons. Using parallel reaction monitoring targeted mass spectrometry we identify the N-terminal amino acids of i-tRNA(AAC)-initiated reporter proteins and show these proteins have altered stability within cells. We also use structural modeling of the peptide deformylase enzyme interaction with position 1 valine peptides to interrogate a potential mechanism for accumulation of formylated-valine proteins observed by mass spectrometry. Our results demonstrate that mutant initiator tRNAs have potential to initiate translation more orthogonally than the native initiator tRNA but their interactions with cellular formyltransferases and peptide deformylases can be inefficient because of the amino acid they are charged with. Additionally, engineered initiator tRNAs may enable tuning of in vivo protein stability through initiation with non-methionine amino acids that alter their interaction with cellular proteases.
Publisher: International Union of Crystallography (IUCr)
Date: 15-05-2013
Publisher: Royal Society of Chemistry (RSC)
Date: 2022
DOI: 10.1039/D2CC04698A
Abstract: Selective O -demethylation of the lignin monoaromatics, syringol and guaiacol, using the peroxygenase activity of two distinct cytochrome P450 enzymes.
Publisher: Proceedings of the National Academy of Sciences
Date: 31-05-2021
Abstract: The structure of single and tandem SHIRT domains from the streptococcal surface protein Sgo_0707 were determined. In conjunction with biophysics and molecular dynamics simulations, the results show that the observed gene length variation would result in differential projection of the host ligand binding domain on the bacterial cell surface. An analysis of long-read DNA sequence data reveals many other repetitive bacterial surface proteins that appear to undergo gene length variation. We propose that these variable-length “Periscope Proteins” represent an important mechanism of bacterial cell surface modification with potential roles in infection and immune evasion.
Publisher: Wiley
Date: 12-2001
DOI: 10.1110/PS.22401
Abstract: Biotin protein ligase of Escherichia coli, the BirA protein, catalyses the covalent attachment of the biotin prosthetic group to a specific lysine of the biotin carboxyl carrier protein (BCCP) subunit of acetyl-CoA carboxylase. BirA also functions to repress the biotin biosynthetic operon and synthesizes its own corepressor, biotinyl-5'-AMP, the catalytic intermediate in the biotinylation reaction. We have previously identified two charge substitution mutants in BCCP, E119K, and E147K that are poorly biotinylated by BirA. Here we used site-directed mutagenesis to investigate residues in BirA that may interact with E119 or E147 in BCCP. None of the complementary charge substitution mutations at selected residues in BirA restored activity to wild-type levels when assayed with our BCCP mutant substrates. However, a BirA variant, in which K277 of the C-terminal domain was substituted with Glu, had significantly higher activity with E119K BCCP than did wild-type BirA. No function has been identified previously for the BirA C-terminal domain, which is distinct from the central domain thought to contain the ATP binding site and is known to contain the biotin binding site. Kinetic analysis of several purified mutant enzymes indicated that a single amino acid substitution within the C-terminal domain (R317E) and located some distance from the presumptive ATP binding site resulted in a 25-fold decrease in the affinity for ATP. Our data indicate that the C-terminal domain of BirA is essential for the catalytic activity of the enzyme and contributes to the interaction with ATP and the protein substrate, the BCCP biotin domain.
Location: United Kingdom of Great Britain and Northern Ireland
Start Date: 2023
End Date: 12-2025
Amount: $522,724.00
Funder: Australian Research Council
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