ARC Centre of Excellence in Synthetic Biology

Despite the environmental persistence of per- and polyfluoroalkyl substances (PFAS), their microbial biotransformation pathways remain poorly characterized due to unidentified metabolites and a lack of direct enzymatic validation. This study examined the roles of acyl-CoA synthetase (ACS) and glutathione S-transferase (GST) from Dietzia aurantiaca strain J3, as well as glutathione (GSH), in the biotransformation of fluorotelomer compounds. Heterologously expressed and purified ACS and GST from strain J3 were evaluated for their catalytic activities toward selected fluorinated and nonfluorinated carboxylates. Results showed that J3 ACS catalyzed the formation of CoA adducts with long-chain fluorinated compounds, including 2H-perfluoro-2-octenoic acid (6:2 FTUCA) and 5:3 fluorotelomer unsaturated carboxylate (5:3 FTUCA), providing the first experimental evidence of CoA adduct formation for eight-carbon PFAS. Notably, no enzymatic activity was observed toward saturated analogues (5:3 FTCA and 6:2 FTCA) or PFOA, with kinetic analysis suggesting that unsaturation and chain length are critical determinants of enzyme reactivity. While GST exhibited no activity toward the tested fluorinated substrates, GSH adducts were observed with 5:3 FTUCA and 6:2 FTUCA via spontaneous, nonenzymatic thiol conjugation. Collectively, these findings support the hypothesis that β-oxidation-like pathways, initiated by CoA conjugation, and GSH-dependent conjugation reactions are both involved in the biotransformation of long-chain polyfluorinated carboxylates.

Polyesters are widely used plastics that persist in the environment due to their resistance to degradation. Microbial polyesterases potentially offer recycling and remediation solutions. However, most polyesterases lack the thermostability and catalytic efficiency required for practical application. Here, we have identified and characterized thermostable bacterial polyesterases from an undercharacterized subfamily of SGNH‐hydrolases (related to PpEST from Pseudomonas oleovorans) and uncovered a previously unreported thermal stabilization mechanism mediated by conformationally flexible N‐terminal regions with features of intrinsically disordered regions. Biochemical assays, structural analysis, small angle X‐ray scattering, X‐ray crystallography, and molecular dynamics simulations suggest that these flexible N‐terminal regions enhance thermal resilience without affecting the catalytic rate or oligomerization in some homologs, while they promote oligomerization and reduce kcat in others. These findings suggest that flexible terminal regions can act as modular stabilizing elements through diverse mechanisms. Our work provides mechanistic insight into an unusual route to protein thermostability, expanding strategies for enzyme engineering, and contributing to the development of robust biocatalysts for polyester degradation.

BackgroundExisting literature has identified inequalities in incidence and health outcomes between urban and rural populations with Alzheimer's disease (AD), however, potential disparities in the diagnostic process are poorly understood. We aimed to investigate differences in the diagnostic pathway – including the completion of key diagnostic investigations and diagnostic wait times – among individuals with all‐cause mild cognitive impairment (MCI) or dementia due to AD residing in rural and urban Australia.MethodWe conducted a cross‐sectional study using data from the Australia Dementia Network (ADNeT) Registry. Patients diagnosed with all‐cause MCI or AD dementia between registry commencement (March 2020) and December 2023 were included. Participants were categorised into three geographic groups – Major Cities (urban), Inner Regional, Outer Regional (both rural) – based on patient postcode (or clinic postcode if unavailable). Logistic and quantile regression models were used to investigate associations between rural/urban residence and the clinical diagnostic pathway.ResultWe identified 3,648 patients, 1,455(39.88%) with all‐cause MCI and 2,193(60.12%) with dementia due to AD. Participants in inner regional areas were more likely (odds ratio [OR]=1.55; 95% confidence interval [CI]=1.14,2.13; p = 0.006) to have had more basic diagnostic investigations completed (including core blood tests, cognitive assessments, functional assessments, structural neuroimaging) compared to those in major cities. However, participants in both inner regional (OR=0.37; 95% CI=0.28,0.48; p <0.001) and outer regional (OR=0.32; 95% CI=0.21,0.48; p <0.001) areas were less likely to have functional neuroimaging completed. Median wait times for an initial appointment following referral to a memory clinic were up to 28 days longer for rural compared to urban participants (Inner regional: Beta (median)=12.92; 95% CI=5.15,20.69; p = 0.001; Outer regional: Beta (median)=27.50; 95% CI=18.80,36.20; p <0.001). However, median wait times from initial appointment to diagnosis were up to 47 days shorter in rural compared to urban residents (Inner regional: Beta (median)=‐46.83; 95% CI=‐52.35,‐41.32; p <0.001; Outer regional: Beta (median)=‐44.42; 95% CI=‐50.35,‐38.49; p <0.001).ConclusionFindings suggest disparities in access to advanced diagnostic investigations and timely initial appointments across rural Australia. These inequalities may preclude access to timely post‐diagnostic services and exacerbate existing barriers to access novel disease modifying therapies which often require advanced diagnostic investigations such as functional neuroimaging.

BackgroundData repositories are critical to advancing open science, ensuring data persistence, and promoting reuse for novel scientific discoveries. In dementia research, platforms such as Dementias Platform Australia, Dementias Platform UK, and the Alzheimer's Disease Data Initiative, have emphasized making data Findable, Accessible, Interoperable, and Reusable (FAIR). However, the decentralized nature of these platforms requires researchers to navigate multiple sites, compile metadata manually, and face challenges in comparing and accessing studies across platforms.MethodTo address these challenges, we developed the Global Ageing and Dementia Study Explorer (GLADSE), an open‐source R Shiny application. The tool collects and standardizes commonly used metadata from partner platforms, including applying the C‐SURV data model themes and domains, to provide rich, detailed and comparable information. The application was built using R Shiny modules under the golem framework, ensuring modularity for scalability and ease of maintenance. GLADSE offers an interactive dashboard with multiple functionalities, including platform and study overviews, filters, visualization plots, and report generation. Additionally, it can be embedded via iframe within partner platform websites, with optional user authentication. A metadata submission tool was developed separately to encourage data custodians to contribute new datasets.ResultGLADSE centralizes metadata exploration, enabling users to identify and compare relevant datasets without navigating multiple platforms. Its modular design allows seamless updates and future expansions. The tool improves data accessibility by featuring diverse metadata and presenting it in an interactive interface. Early feedback highlights its effectiveness in improving data discovery and enabling cross‐platform collaboration, including federated analysis. By integrating global resources, the tool maximizes the utility of existing research infrastructure and facilitates global partnerships.ConclusionGLADSE advances dementia research making data FAIR and fostering collaboration across dementia platforms. Planned future developments, such as incorporating measurement‐level metadata standardised using the C‐SURV ontology will enhance the utility of study filters and visualization plots. Expanding to include detailed imaging and genomic metadata will further strengthen its utility. By proposing a solution to the decentralized data platforms and promoting global collaboration, GLADSE maximizes the value of public investments in research infrastructures, supports critical progress in understanding, preventing, and treating dementia, a pressing public health challenge.

Integrons are genetic elements that facilitate gene acquisition. They have been extensively studied in clinical bacteria, but their evolutionary role in phytopathogens remains underexplored. Here, we analysed complete genomes of Xanthomonas species to investigate the origin, distribution and functional dynamics of integrons in this genus. We found that 93% of genomes harboured integrons. The integron-integrase gene intI was predominantly located downstream of ilvD, indicating an ancestral acquisition of integrons, predating diversification within the genus. Phylogenetic analyses support vertical inheritance of intI, with the exception of rare horizontal gene transfer events, notably in Xanthomonas arboricola. Despite their widespread presence, full-length intI genes and active integron platforms are only retained in some species, especially Xanthomonas campestris, which shows high integron gene cassette variability and functional integron activity. In contrast, species such as Xanthomonas cissicola and Xanthomonas phaseoli exhibit widespread intI inactivation, likely occurring early in their divergence, leading to more stable cassette arrays and conserved integron-associated phenotypes. The number and diversity of genes within cassette arrays varied significantly by species and, to a lesser extent, by the ecological context of plant host cultivation. While most cassettes encoded proteins without a known function, those with annotated roles were associated with stress response mechanism, competitive exclusion and plant-associated functions. Together, our findings demonstrate that integrons in Xanthomonas likely originated from a single ancient acquisition event, preceding genus-wide speciation, and have co-evolved with Xanthomonas pathovars as they adapted to distinct plant hosts.

Mutations in the TANGO2 gene cause an autosomal recessive disorder characterised by developmental delay, stress-induced episodic rhabdomyolysis, and cardiac arrhythmias along with severe metabolic crises. Although TANGO2 mutations result in a well characterised disease pathology, the function of TANGO2 is still unknown. To investigate the function of TANGO2, we knocked out the TANGO2 gene in human cells and mice. We identify that loss of TANGO2 impairs intermediate filament structure, resulting in fragmented mitochondrial networks and formation of cup-like mitochondria. In male mice, loss of TANGO2 caused heart defects, reduced muscle function and glucose intolerance by remodelling of intermediate filaments, which altered the mitochondrial and cytoplasmic proteomes, N-glycosylation and nucleocytoplasmic O-GlcNAcylation. We identify that TANGO2 binds the small heat shock protein crystallin alpha B (CRYAB) to prevent the aggregation of the intermediate filament desmin and in the absence of TANGO2, mice develop desminopathy, which is consistent with features found in patients carrying mutations in either desmin or CRYAB.