Bio21 - Research Publications

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    NMR techniques for investigating antimicrobial peptides in model membranes and bacterial cells
    Sani, M-A ; Rajput, S ; Keizer, DW ; Separovic, F (Elsevier BV, 2024-04)
    AMPs are short, mainly cationic membrane-active peptides found in all living organism. They perform diverse roles including signaling and acting as a line of defense against bacterial infections. AMPs have been extensively investigated as templates to facilitate the development of novel antimicrobial therapeutics. Understanding the interplay between these membrane-active peptides and the lipid membranes is considered to be a significant step in elucidating the specific mechanism of action of AMPs against prokaryotic and eukaryotic cells to aid the development of new therapeutics. In this review, we have provided a brief overview of various NMR techniques commonly used for studying AMP structure and AMP-membrane interactions in model membranes and whole cells.
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    Development of Matrix-Embedded Bovine Tracheal Organoids to Study the Innate Immune Response against Bovine Respiratory Disease
    Quah, PS ; Tran, BM ; Corbin, VDA ; Chang, JJ-Y ; Wong, CY ; Diaz-Méndez, A ; Hartley, CA ; Zeng, W ; Hanssen, E ; Trifunovic, Z ; Reading, PC ; Jackson, DC ; Vincan, E ; Coin, LJM ; Deliyannis, G (MDPI, 2023)
    Bovine respiratory disease (BRD) is the leading cause of morbidity and mortality in feedlot cattle. Bovine herpesvirus-1 (BHV-1) is one of the main culprits of BRD; however, research on BHV-1 is hampered by the lack of suitable models for infection and drug testing. In this study, we established a novel bovine tracheal organoid culture grown in a basement membrane extract type 2 (BME2) matrix and compared it with the air–liquid interface (ALI) culture system. After differentiation, the matrix-embedded organoids developed beating cilia and demonstrated a transcriptomic profile similar to the ALI culture system. The matrix-embedded organoids were also highly susceptible to BHV-1 infection and immune stimulation by Pam2Cys, an immunomodulator, which resulted in robust cytokine production and tracheal antimicrobial peptide mRNA upregulation. However, treatment of bovine tracheal organoid cultures with Pam2Cys was not sufficient to inhibit viral infection or replication, suggesting a role of the non-epithelial cellular microenvironment in vivo.
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    The Proteome and Lipidome of Extracellular Vesicles from Haemonchus contortus to Underpin Explorations of Host-Parasite Cross-Talk
    Wang, T ; Koukoulis, TF ; Vella, LJ ; Su, H ; Purnianto, A ; Nie, S ; Ang, C-S ; Ma, G ; Korhonen, PK ; Taki, AC ; Williamson, NA ; Reid, GE ; Gasser, RB (MDPI, 2023-07)
    Many parasitic worms have a major adverse impact on human and animal populations worldwide due to the chronicity of their infections. There is a growing body of evidence indicating that extracellular vesicles (EVs) are intimately involved in modulating (suppressing) inflammatory/immune host responses and parasitism. As one of the most pathogenic nematodes of livestock animals, Haemonchus contortus is an ideal model system for EV exploration. Here, employing a multi-step enrichment process (in vitro culture, followed by ultracentrifugation, size exclusion and filtration), we enriched EVs from H. contortus and undertook the first comprehensive (qualitative and quantitative) multi-omic investigation of EV proteins and lipids using advanced liquid chromatography-mass spectrometry and informatics methods. We identified and quantified 561 proteins and 446 lipids in EVs and compared these molecules with those of adult worms. We identified unique molecules in EVs, such as proteins linked to lipid transportation and lipid species (i.e., sphingolipids) associated with signalling, indicating the involvement of these molecules in parasite-host cross-talk. This work provides a solid starting point to explore the functional roles of EV-specific proteins and lipids in modulating parasite-host cross-talk, and the prospect of finding ways of disrupting or interrupting this relationship to suppress or eliminate parasite infection.
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    Selenium Nanoparticles as Potential Drug-Delivery Systems for the Treatment of Parkinson's Disease
    Kalcec, N ; Peranic, N ; Mamic, I ; Beus, M ; Hall, CR ; Smith, TA ; Sani, MA ; Turcic, P ; Separovic, F ; Vrcek, IV (AMER CHEMICAL SOC, 2023-09-20)
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    Encounter Complexes Between the N-terminal of Neurotensin with the Extracellular Loop 2 of the Neurotensin Receptor 1 Steer Neurotensin to the Orthosteric Binding Pocket
    Asadollahi, K ; Rajput, S ; Jameson, GNL ; Scott, DJ ; Gooley, PR (ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 2023-10-15)
    Neurotensin (NT) is a linear disordered peptide that activates two different class A GPCRs, neurotensin receptor 1 (NTS1) and NTS2. Resolved structures of the complex of the C-terminal fragment of NT, NT8-13, with NTS1 shows the peptide takes a well-defined structure in the bound state. However, the mechanisms underlying NT recognition of NTS1, and the conformational transition of NT upon binding NTS1 is an open question that if answered may aid discovery of highly selective drugs and reveal potential secondary binding sites on the surface of the receptor. Herein we investigated the interactions guiding NT to the orthosteric binding pocket of NTS1 by combining NMR experiments with kinetic analysis of the binding pathway using stopped-flow fluorescence and mutagenesis on both NT and NTS1. We show the presence of transient structures in the middle part of NT that kinetically regulate the binding of NT to NTS1. Moreover, our results indicate that the binding pathway of NT onto NTS1 is mediated via electrostatic interactions between the N-terminal region of NT with the extracellular loop 2 of NTS1. These interactions induce backbone conformational changes in neurotensin similar to the bound-state neurotensin, suggesting that the N-terminal region of NT and these interactions should be considered for development of selective drugs against NTS1.
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    Plasma lipids are dysregulated preceding diagnosis of preeclampsia or delivery of a growth restricted infant
    Bartho, LA ; Keenan, E ; Walker, SP ; MacDonald, TM ; Nijagal, B ; Tong, S ; Kaitu'u-Lino, TJ (ELSEVIER, 2023-08)
    BACKGROUND: Lipids serve as multifunctional metabolites that have important implications for the pregnant mother and developing fetus. Abnormalities in lipids have emerged as potential risk factors for pregnancy diseases, such as preeclampsia and fetal growth restriction. The aim of this study was to assess the potential of lipid metabolites for detection of late-onset preeclampsia and fetal growth restriction. METHODS: We used a case-cohort of 144 maternal plasma samples at 36 weeks' gestation from patients before the diagnosis of late-onset preeclampsia (n = 22), delivery of a fetal growth restricted infant (n = 55, defined as <5th birthweight centile), gestation-matched controls (n = 72). We performed liquid chromatography-tandem mass spectrometry (LC-QQQ) -based targeted lipidomics to identify 421 lipids, and fitted logistic regression models for each lipid, correcting for maternal age, BMI, smoking, and gestational diabetes. FINDINGS: Phosphatidylinositol 32:1 (AUC = 0.81) and cholesterol ester 17:1 (AUC = 0.71) best predicted the risk of developing preeclampsia or delivering a fetal growth restricted infant, respectively. Five times repeated five-fold cross validation demonstrated the lipids alone did not out-perform existing protein biomarkers, soluble tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF) for the prediction of preeclampsia or fetal growth restriction. However, lipids combined with sFlt-1 and PlGF measurements improved disease prediction. INTERPRETATION: This study successfully identified 421 lipids in maternal plasma collected at 36 weeks' gestation from participants who later developed preeclampsia or delivered a fetal growth restricted infant. Our results suggest the predictive capacity of lipid measurements for gestational disorders holds the potential to improve non-invasive assessment of maternal and fetal health. FUNDING: This study was funded by a grant from National Health and Medical Research Council.
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    A sandwich-like structural model revealed for quasi-2D perovskite films
    Zheng, F ; Hall, CR ; Angmo, D ; Zuo, C ; Rubanov, S ; Wen, Z ; Bradley, SJ ; Hao, X-T ; Gao, M ; Smith, TA ; Ghiggino, KP (Royal Society of Chemistry, 2021-04-28)
    The excellent performance and stability of perovskite solar cells (PSCs) based on quasi-2D Ruddlesden–Popper perovskites (RPPs) holds promise for their commercialization. Further improvement in the performance of 2D PSCs requires a detailed understanding of the microstructure of the quasi-2D perovskite films. Based on scanning transmission electron microscopy (STEM), time-resolved photoluminescence, and transient absorption measurements, a new sandwich-like structural model is proposed to describe the phase distribution of RPPs. In contrast to the conventional gradient distribution, it is found that small-n RPPs are sandwiched between large-n RPP phase layers at the front and back sides owing to crystallization initiated from both interfaces during film formation. This sandwich-like distribution profile facilitates excitons funneling from the film interior to both surfaces for dissociation while free carriers transport via large-n channels that permeate the film to ensure efficient charge collection by the corresponding electrodes, which is favorable for high-performance photovoltaics. This discovery provides a new fundamental understanding of the operating principles of 2D PSCs and has valuable implications for the design and optimization strategies of optoelectronic devices based on quasi-2D RPPs films.
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    Supramolecular Assembly of Polyphenols and Nucleic Acids by Thermal Cycling for Immune Cell Activation
    Qu, Y ; Zhu, H ; Lin, Z ; Vanni, D ; Bhangu, SK ; Dyett, B ; Sani, M-A ; Cortez-Jugo, C ; Caruso, F ; Cavalieri, F (American Chemical Society (ACS), 2023)
    Supramolecular assembly of polyphenols and biomacromolecules (proteins and nucleic acids) has emerged as a versatile and simple strategy to construct nanomaterials with biological activity. Here, we report a strategy to finely control the supramolecular assembly of tannic acid and oligonucleotides into uniform and stable nanoparticles by exploiting the thermal cycling of tannic acid. The equilibrium of complexation is investigated, and individual nanoparticles are resolved with nanoscale resolution by using stochastic optical reconstruction microscopy. The nanoparticles incorporating cytosine phosphoguanine (CpG) oligonucleotides are efficiently taken up by cells and trafficked via endo/lysosomal compartments and induce up to a 7-fold increase in tumor necrosis factor secretion in RAW 264.7 macrophage cells compared with naked CpG oligonucleotides. This work highlights the potential of this simple approach to engineer two-component tannic acid–oligonucleotide nanoparticles for the intracellular delivery of therapeutic nucleic acids.
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    Characterising the influence of milk fat towards an application for extrusion-based 3D-printing of casein-whey protein suspensions via the pH-temperature-route
    Daffner, K ; Ong, L ; Hanssen, E ; Gras, S ; Mills, T (ELSEVIER SCI LTD, 2021-09)
    This study presents the design and characterisation of casein−whey protein suspensions (8.0/10.0% (w/w) casein and 2.0/2.5% (w/w) whey protein) mixed with dairy fat (1.0, 2.5 and 5.0% (w/w) total fat) processed via the pH−temperature-route in preparation for 3D-printing. Mechanical treatment was applied to significantly decrease the particle size of the milk fat globules and increase surface area, creating small fat globules (<1 μm) covered with proteins, which could act as pseudo protein particles during gelation. Different proteins covered the fat globule surface after mechanical treatment, as a result of differences in the pH adjusted just prior to heating (6.55, 6.9 or 7.1). The protein-fat suspensions appeared similar by transmission electron cryogenic microscopy and the zeta-potential of all particles was unchanged by the heating pH, with a similar charge to the solution (~−20 mV) occurring after acidification (pH 4.8/5.0) at low temperatures (2 °C). A low heating pH (6.55) resulted in increased sol−gel transition temperatures (G՛ = 1 Pa) and a decreased rate of aggregation for protein−fat suspensions. A higher heating pH (6.9 and 7.1) caused an increased rate of aggregation (aggregation rate ≥ 250 Pa/10 K), resulting in materials more promising for application in extrusion-based printing. 3D-printing of formulations into small rectangles, inclusive of a sol−gel transition in a heated nozzle, was conducted to relate the aggregation rate towards printability.
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    Brownian Tree‐Shaped Dendrites in Quasi‐2D Perovskite Films and Their Impact on Photovoltaic Performance
    Zheng, F ; Angmo, D ; Hall, CR ; Rubanov, S ; Yuan, F ; Laird, JS ; Gao, M ; Smith, TA ; Ghiggino, KP (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.