ORCID Profile
0000-0003-0989-7734
Current Organisations
University of Adelaide
,
Walter and Eliza Hall Institute of Medical Research
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Publisher: International Union of Crystallography (IUCr)
Date: 28-04-2021
DOI: 10.1107/S2053230X21004052
Abstract: The phase problem is a persistent bottleneck that impedes the structure-determination pipeline and must be solved to obtain atomic resolution crystal structures of macromolecules. Although molecular replacement has become the predominant method of solving the phase problem, many scenarios still exist in which experimental phasing is needed. Here, a proof-of-concept study is presented that shows the efficacy of using tetrabromoterephthalic acid (B4C) as an experimental phasing compound. Incorporating B4C into the crystal lattice using co-crystallization, the crystal structure of hen egg-white lysozyme was solved using MAD phasing. The strong anomalous signal generated by its four Br atoms coupled with its compatibility with commonly used crystallization reagents render B4C an effective experimental phasing compound that can be used to overcome the phase problem.
Publisher: International Union of Crystallography (IUCr)
Date: 26-01-2022
DOI: 10.1107/S2059798321012031
Abstract: Purine biosynthesis is a fundamental cellular process that sustains life by maintaining the intracellular pool of purines for DNA/RNA synthesis and signal transduction. As an integral determinant of fungal survival and virulence, the enzymes in this metabolic pathway have been pursued as potential antifungal targets. Guanosine monophosphate (GMP) synthase has been identified as an attractive target as it is essential for virulence in the clinically prominent fungal pathogens Aspergillus fumigatus , Candida albicans and Cryptococcus neoformans . However, a lack of structural information on GMP synthase has hindered drug-design efforts. Here, the first structure of a GMP synthase of fungal origin, that from A. fumigatus (at 2.3 Å resolution), is presented. Structural analysis of GMP synthase shows a distinct absence of the D1 dimerization domain that is present in the human homologue. Interestingly, A. fumigatus GMP synthase adopts a dimeric state, as determined by native mass spectrometry and gel-filtration chromatography, in contrast to the monomeric human homologue. Analysis of the substrate-binding pockets of A. fumigatus GMP synthase reveals key differences in the ATP- and XMP-binding sites that can be exploited for species-specific inhibitor drug design. Furthermore, the inhibitory activities of the glutamine analogues acivicin (IC 50 = 16.6 ± 2.4 µ M ) and 6-diazo-5-oxo-L-norleucine (IC 50 = 29.6 ± 5.6 µ M ) against A. fumigatus GMP synthase are demonstrated. Together, these data provide crucial structural information required for specifically targeting A. fumigatus GMP synthase for future antifungal drug-discovery endeavours.
Publisher: Wiley
Date: 22-03-2022
DOI: 10.1002/PROT.26331
Abstract: The metabolic enzyme, enolase, plays a crucial role in the cytoplasm where it maintains cellular energy production within the process of glycolysis. The main role of enolase in glycolysis is to convert 2‐phosphoglycerate to phosphoenolpyruvate however, enolase can fulfill roles that deviate from this function. In pathogenic bacteria and fungi, enolase is also located on the cell surface where it functions as a virulence factor. Surface‐expressed enolase is a receptor for human plasma proteins, including plasminogen, and this interaction facilitates nutrient acquisition and tissue invasion. A novel approach to developing antifungal drugs is to inhibit the formation of this complex. To better understand the structure of enolase and the interactions that may govern complex formation, we have solved the first X‐ray crystal structure of enolase from Aspergillus fumigatus (2.0 Å) and have shown that it preferentially adopts a dimeric quaternary structure using native mass spectrometry. Two additional X‐ray crystal structures of A. fumigatus enolase bound to the endogenous substrate 2‐phosphoglycerate and product phosphoenolpyruvate were determined and kinetic characterization was carried out to better understand the details of its canonical function. From these data, we have produced a model of the A. fumigatus enolase and human plasminogen complex to provide structural insights into the mechanisms of virulence and aid future development of small molecules or peptidomimetics for antifungal drug design.
Publisher: Frontiers Media SA
Date: 12-01-2021
DOI: 10.3389/FMOLB.2020.621366
Abstract: The impact of invasive fungal infections on human health is a serious, but largely overlooked, public health issue. Commonly affecting the immunocompromised community, fungal infections are predominantly caused by species of Candida, Cryptococcus , and Aspergillus . Treatments are reliant on the aggressive use of pre-existing antifungal drug classes that target the fungal cell wall and membrane. Despite their frequent use, these drugs are subject to unfavorable drug-drug interactions, can cause undesirable side-effects and have compromised efficacy due to the emergence of antifungal resistance. Hence, there is a clear need to develop novel classes of antifungal drugs. A promising approach involves exploiting the metabolic needs of fungi by targeted interruption of essential metabolic pathways. This review highlights potential antifungal targets including enolase, a component of the enolase-plasminogen complex, and enzymes from the mannitol biosynthesis and purine nucleotide biosynthesis pathways. There has been increased interest in the enzymes that comprise these particular pathways and further investigation into their merits as antifungal targets and roles in fungal survival and virulence are warranted. Disruption of these vital processes by targeting unconventional pathways with small molecules or antibodies may serve as a promising approach to discovering novel classes of antifungals.
Publisher: Wiley
Date: 12-01-2021
DOI: 10.1002/MGG3.1593
Abstract: Vanishing white matter (VWM) is a leukodystrophy, caused by recessive mutations in eukaryotic initiation factor 2B (eIF2B)‐subunit genes ( EIF2B1–EIF2B5 ) 80% are missense mutations. Clinical severity is highly variable, with a strong, unexplained genotype–phenotype correlation. With information from a recent natural history study, we severity‐graded 97 missense mutations. Using in silico modeling, we created a new human eIF2B model structure, onto which we mapped the missense mutations. Mutated residues were assessed for location in subunits, eIF2B complex, and functional domains, and for information on biochemical activity. Over 50% of mutations have (ultra‐)severe phenotypic effects. About 60% affect the ε‐subunit, containing the catalytic domain, mostly with (ultra‐)severe effects. About 55% affect subunit cores, with variable clinical severity. About 36% affect subunit interfaces, mostly with severe effects. Very few mutations occur on the external eIf2B surface, perhaps because they have minor functional effects and are tolerated. One external surface mutation affects eIF2B‐substrate interaction and is associated with ultra‐severe phenotype. Mutations that lead to (ultra‐)severe disease mostly affect amino acids with pivotal roles in complex formation and function of eIF2B. Therapies for VWM are emerging and reliable mutation‐based phenotype prediction is required for propensity score matching for trials and in the future for in idualized therapy decisions.
Publisher: Cold Spring Harbor Laboratory
Date: 11-04-2020
DOI: 10.1101/2020.04.09.034942
Abstract: Monitoring the mutation dynamics of SARS-CoV-2 is critical for the development of effective approaches to contain the pathogen. By analyzing 106 SARS-CoV-2 and 39 SARS genome sequences, we provided direct genetic evidence that SARS-CoV-2 has a much lower mutation rate than SARS. Minimum Evolution phylogeny analysis revealed the putative original status of SARS-CoV-2 and the early-stage spread history. The discrepant phylogenies for the spike protein and its receptor binding domain proved a previously reported structural rearrangement prior to the emergence of SARS-CoV-2. Despite that we found the spike glycoprotein of SARS-CoV-2 is particularly more conserved, we identified a receptor binding domain mutation that leads to weaker ACE2 binding capability based on in silico simulation, which concerns a SARS-CoV-2 s le collected on 27 th January 2020 from India. This represents the first report of a significant SARS-CoV-2 mutant, and requires attention from researchers working on vaccine development around the world. Based on the currently available genome sequence data, we provided direct genetic evidence that the SARS-COV-2 genome has a much lower mutation rate and genetic ersity than SARS during the 2002-2003 outbreak. The spike (S) protein encoding gene of SARS-COV-2 is found relatively more conserved than other protein-encoding genes, which is a good indication for the ongoing antiviral drug and vaccine development. Minimum Evolution phylogeny analysis revealed the putative original status of SARS-CoV-2 and the early-stage spread history. We confirmed a previously reported rearrangement in the S protein arrangement of SARS-COV-2, and propose that this rearrangement should have occurred between human SARS-CoV and a bat SARS-CoV, at a time point much earlier before SARS-COV-2 transmission to human. We provided first evidence that a mutated SARS-COV-2 with reduced human ACE2 receptor binding affinity have emerged in India based on a s le collected on 27th January 2020.
Publisher: MyJove Corporation
Date: 16-01-2021
DOI: 10.3791/61894
Publisher: Wiley
Date: 06-11-2020
DOI: 10.1111/FEBS.15607
Publisher: Elsevier BV
Date: 09-2020
Location: Australia
No related grants have been discovered for Stephanie Nguyen.