School of Chemistry - Research Publications
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ItemDiscovery and Biosynthesis of the Cytotoxic Polyene Terpenomycin in Human Pathogenic Nocardia(AMER CHEMICAL SOC, 2023-07-27)Nocardia are opportunistic human pathogens that can cause a range of debilitating and difficult to treat infections of the lungs, brain, skin, and soft tissues. Despite their close relationship to the well-known secondary metabolite-producing genus, Streptomyces, comparatively few natural products are known from the Nocardia, and even less is known about their involvement in the pathogenesis. Here, we combine chemistry, genomics, and molecular microbiology to reveal the production of terpenomycin, a new cytotoxic and antifungal polyene from a human pathogenic Nocardia terpenica isolate. We unveil the polyketide synthase (PKS) responsible for terpenomycin biosynthesis and show that it combines several unusual features, including "split", skipped, and iteratively used modules, and the use of the unusual extender unit methoxymalonate as a starter unit. To link genes to molecules, we constructed a transposon mutant library in N. terpenica, identifying a terpenomycin-null mutant with an inactivated terpenomycin PKS. Our findings show that the neglected actinomycetes have an unappreciated capacity for the production of bioactive molecules with unique biosynthetic pathways waiting to be uncovered and highlights these organisms as producers of diverse natural products.
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ItemBrownian Tree‐Shaped Dendrites in Quasi‐2D Perovskite Films and Their Impact on Photovoltaic Performance(Wiley, 2022-05)Quasi-2D Ruddlesden–Popper perovskites (RPPs) are candidates for constructing perovskite solar cells (PSCs) with superior stability due to their tolerance to the external environment. Fully understanding the film growth mechanism and structure is crucial to further improve the performance of 2D-PSCs while maintaining device stability. In this work, the origin of Brownian tree-shaped dendrites formed in hot-cast methylammonium chloride (MACl)-doped BA2MAn−1PbnI3n+1 ( = 5) quasi-2D perovskite films are reported. Investigations based on optical, electronic, atomic force, and fluorescence microscopies reveal that the dendrites are assembled from large-n RPPs-dominated grains, while the nondendritic film area is composed of small-n RPPs grains and associated with film surface pits caused by the evaporation of MACl. It is proposed that these dendrites are grown by the diffusion-limited aggregation of the MA-rich intermediate phase domains that initially crystallize from the precursor. The formation of these dendrites in quasi-2D perovskite films upon MACl doping is accompanied by improved organization and crystallinity of the 2D RPPs, which benefits the photovoltaic performance. This work provides new insights into the formation mechanism of quasi-2D perovskite films that should assist device engineering strategies to further improve the performance of 2D PSCs.
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ItemNo Preview AvailableIncorporating whey protein aggregates produced with heat and ultrasound treatment into rennet gels and model non-fat cheese systems(Elsevier, 2020-12-01)Native whey proteins (WP) are expulsed from cheese coagulation during syneresis. Although incorporating denatured WP aggregates into cheese gels has been previously proposed to improve the overall cheese yield, the effects of WP aggregate properties on gelation kinetics and protein retention are not yet fully understood. In this study, heat and power ultrasound were used to produce denatured whey protein aggregates with a wide range of sizes. The effects of size and hydrophobicity differences in WP aggregates produced by heat and heat coupled with ultrasound were investigated in relation to the kinetics of rennet gelation and protein retention in model non-fat cheddar cheeses. Rheological measurements showed that sufficiently large, denatured WP aggregates could avoid impairment of rennet gelation caused by native WP, irrespective of changes in the soluble calcium concentration or the surface hydrophobicity of the aggregates. WP aggregates formed by the combined heat and ultrasound treatment were more hydrophobic than the larger heat-treated aggregates and were better retained in the cheese. However, inclusion of sufficiently large aggregates in cheese milk conferred an openness to the cheese microstructure and showed promise in improving the otherwise rigid non-fat cheese.
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ItemEditors’ page - Historical Records of Australian Science(CSIRO, 2023-07-24)
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ItemFamilial ALS-associated SFPQ variants promote the formation of SFPQ cytoplasmic aggregates in primary neurons(ROYAL SOC, 2022-09-28)Splicing factor proline- and glutamine-rich (SFPQ) is a nuclear RNA-binding protein that is involved in a wide range of physiological processes including neuronal development and homeostasis. However, the mislocalization and cytoplasmic aggregation of SFPQ are associated with the pathophysiology of amyotrophic lateral sclerosis (ALS). We have previously reported that zinc mediates SFPQ polymerization and promotes the formation of cytoplasmic aggregates in neurons. Here we characterize two familial ALS (fALS)-associated SFPQ variants, which cause amino acid substitutions in the proximity of the SFPQ zinc-coordinating centre (N533H and L534I). Both mutants display increased zinc-binding affinities, which can be explained by the presence of a second zinc-binding site revealed by the 1.83 Å crystal structure of the human SFPQ L534I mutant. Overexpression of these fALS-associated mutants significantly increases the number of SFPQ cytoplasmic aggregates in primary neurons. Although they do not affect the density of dendritic spines, the presence of SFPQ cytoplasmic aggregates causes a marked reduction in the levels of the GluA1, but not the GluA2 subunit of AMPA-type glutamate receptors on the neuronal surface. Taken together, our data demonstrate that fALS-associated mutations enhance the propensity of SFPQ to bind zinc and form aggregates, leading to the dysregulation of AMPA receptor subunit composition, which may contribute to neuronal dysfunction in ALS.
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ItemParaspeckle subnuclear bodies depend on dynamic heterodimerisation of DBHS RNA-binding proteins via their structured domains(ELSEVIER, 2022-11-04)RNA-binding proteins of the DBHS (Drosophila Behavior Human Splicing) family, NONO, SFPQ, and PSPC1 have numerous roles in genome stability and transcriptional and posttranscriptional regulation. Critical to DBHS activity is their recruitment to distinct subnuclear locations, for example, paraspeckle condensates, where DBHS proteins bind to the long noncoding RNA NEAT1 in the first essential step in paraspeckle formation. To carry out their diverse roles, DBHS proteins form homodimers and heterodimers, but how this dimerization influences DBHS localization and function is unknown. Here, we present an inducible GFP-NONO stable cell line and use it for live-cell 3D-structured illumination microscopy, revealing paraspeckles with dynamic, twisted elongated structures. Using siRNA knockdowns, we show these labeled paraspeckles consist of GFP-NONO/endogenous SFPQ dimers and that GFP-NONO localization to paraspeckles depends on endogenous SFPQ. Using purified proteins, we confirm that partner swapping between NONO and SFPQ occurs readily in vitro. Crystallographic analysis of the NONO-SFPQ heterodimer reveals conformational differences to the other DBHS dimer structures, which may contribute to partner preference, RNA specificity, and subnuclear localization. Thus overall, our study suggests heterodimer partner availability is crucial for NONO subnuclear distribution and helps explain the complexity of both DBHS protein and paraspeckle dynamics through imaging and structural approaches.
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ItemGenetic and chemical validation of Plasmodium falciparum aminopeptidase PfA-M17 as a drug target in the hemoglobin digestion pathway(eLIFE SCIENCES PUBL LTD, 2022-09-13)Plasmodium falciparum, the causative agent of malaria, remains a global health threat as parasites continue to develop resistance to antimalarial drugs used throughout the world. Accordingly, drugs with novel modes of action are desperately required to combat malaria. P. falciparum parasites infect human red blood cells where they digest the host's main protein constituent, hemoglobin. Leucine aminopeptidase PfA-M17 is one of several aminopeptidases that have been implicated in the last step of this digestive pathway. Here, we use both reverse genetics and a compound specifically designed to inhibit the activity of PfA-M17 to show that PfA-M17 is essential for P. falciparum survival as it provides parasites with free amino acids for growth, many of which are highly likely to originate from hemoglobin. We further show that loss of PfA-M17 results in parasites exhibiting multiple digestive vacuoles at the trophozoite stage. In contrast to other hemoglobin-degrading proteases that have overlapping redundant functions, we validate PfA-M17 as a potential novel drug target.
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ItemAlkyne Activation in the Diversity Oriented Synthesis of sp(2)-Rich Scaffolds: A Biased Library Approach for Targeting Polynucleotides (DNA/RNA)(WILEY-V C H VERLAG GMBH, 2022-11-03)Polynucleotides, DNA and RNA (mRNA and non-coding RNAs) are critically involved in the molecular pathways of disease. Small molecule binding interactions with polynucleotides can modify functional polynucleotide topologies and/or their interactions with proteins. Current approaches to library design (lead-like or fragment-like libraries) are based on protein-ligand interactions and often include careful consideration of the 3-dimensional orientation of binding motifs and exclude π-rich compounds (polyfused aromatics) to avoid off-target R/DNA interactions. In contrast to proteins, where π,π-interactions are weak, polynucleotides can form strong π,π-interactions with suitable π-rich ligands. To assist in designing a polynucleotide-biased library, a scaffold-divergent synthesis approach to polyfused aromatic scaffolds has been undertaken. Initial screening hits that form moderately stable polynucleotide-ligand-protein ternary complexes can be further optimized through judicious incorporation of substituents on the scaffold to increase protein-ligand interactions. An example of this approach is given for topoisomerase-1 (TOP1), generating a novel TOP1 inhibitory chemotype.
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ItemCytochrome P450Blt Enables Versatile Peptide Cyclisation to Generate Histidine- and Tyrosine-Containing Crosslinked Tripeptide Building Blocks.(Wiley, 2022-09-12)We report our investigation of the utility of peptide crosslinking cytochrome P450 enzymes from biarylitide biosynthesis to generate a range of cyclic tripeptides from simple synthons. The crosslinked tripeptides produced by this P450 include both tyrosine-histidine (A-N-B) and tyrosine-tryptophan (A-O-B) crosslinked tripeptides, the latter a rare example of a phenolic crosslink to an indole moiety. Tripeptides are easily isolated following proteolytic removal of the leader peptide and can incorporate a wide range of amino acids in the residue inside the crosslinked tripeptide. Given the utility of peptide crosslinks in important natural products and the synthetic challenge that these can represent, P450 enzymes have the potential to play roles as important tools in the generation of high-value cyclic tripeptides for incorporation in synthesis, which can be yet further diversified using selective chemical techniques through specific handles contained within these tripeptides.