ORCID Profile
0000-0001-7999-1385
Current Organisation
University of Adelaide
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Publisher: American Chemical Society (ACS)
Date: 21-02-2023
Publisher: Elsevier BV
Date: 10-2022
Publisher: Elsevier BV
Date: 03-2020
Publisher: Wiley
Date: 26-07-2021
DOI: 10.1111/FEBS.16122
Abstract: Under certain cellular conditions, functional proteins undergo misfolding, leading to a transition into oligomers which precede the formation of amyloid fibrils. Misfolding proteins are associated with neurodegenerative diseases such as Alzheimer’s and Parkinson’s diseases. While the importance of lipid membranes in misfolding and disease aetiology is broadly accepted, the influence of lipid membranes during therapeutic design has been largely overlooked. This study utilized a biophysical approach to provide mechanistic insights into the effects of two lipid membrane systems (anionic and zwitterionic) on the inhibition of amyloid‐β 40 and α‐synuclein amyloid formation at the monomer, oligomer and fibril level. Large unilamellar vesicles (LUVs) were shown to increase fibrillization and largely decrease the effectiveness of two well‐known polyphenol fibril inhibitors, (‐)‐epigallocatechin gallate (EGCG) and resveratrol however, use of immunoblotting and ion mobility mass spectrometry revealed this occurs through varying mechanisms. Oligomeric populations in particular were differentially affected by LUVs in the presence of resveratrol, an elongation phase inhibitor, compared to EGCG, a nucleation targeted inhibitor. Ion mobility mass spectrometry showed EGCG interacts with or induces more compact forms of monomeric protein typical of off‐pathway structures however, binding is reduced in the presence of LUVs, likely due to partitioning in the membrane environment. Competing effects of the lipids and inhibitor, along with reduced inhibitor binding in the presence of LUVs, provide a mechanistic understanding of decreased inhibitor efficacy in a lipid environment. Together, this study highlights that amyloid inhibitor design may be misguided if effects of lipid membrane composition and architecture are not considered during development.
Publisher: American Chemical Society (ACS)
Date: 06-02-2023
Publisher: Portland Press Ltd.
Date: 10-06-2020
DOI: 10.1042/BCJ20200290
Abstract: Amyloid beta peptide (Aβ42) aggregation in the brain is thought to be responsible for the onset of Alzheimer's disease, an insidious condition without an effective treatment or cure. Hence, a strategy to prevent aggregation and subsequent toxicity is crucial. Bio-inspired peptide-based molecules are ideal candidates for the inhibition of Aβ42 aggregation, and are currently deemed to be a promising option for drug design. In this study, a hexapeptide containing a self-recognition component unique to Aβ42 was designed to mimic the β-strand hydrophobic core region of the Aβ peptide. The peptide is comprised exclusively of D-amino acids to enhance specificity towards Aβ42, in conjunction with a C-terminal disruption element to block the recruitment of Aβ42 monomers on to fibrils. The peptide was rationally designed to exploit the synergy between the recognition and disruption components, and incorporates features such as hydrophobicity, β-sheet propensity, and charge, that all play a critical role in the aggregation process. Fluorescence assays, native ion-mobility mass spectrometry (IM-MS) and cell viability assays were used to demonstrate that the peptide interacts with Aβ42 monomers and oligomers with high specificity, leading to almost complete inhibition of fibril formation, with essentially no cytotoxic effects. These data define the peptide-based inhibitor as a potentially potent anti-amyloid drug candidate for this hitherto incurable disease.
Publisher: Springer Science and Business Media LLC
Date: 08-03-2018
Publisher: Elsevier BV
Date: 02-2015
DOI: 10.1016/J.CHEMBIOL.2015.01.001
Abstract: Serine phosphorylation of the mammalian small heat-shock protein Hsp27 at residues 15, 78, and 82 is thought to regulate its structure and chaperone function however, the site-specific impact has not been established. We used mass spectrometry to assess the combinatorial effect of mutations that mimic phosphorylation upon the oligomeric state of Hsp27. Comprehensive dimerization yielded a relatively uncrowded spectrum, composed solely of even-sized oligomers. Modification at one or two serines decreased the average oligomeric size, while the triple mutant was predominantly a dimer. These changes were reflected in a greater propensity for oligomers to dissociate upon increased modification. The ability of Hsp27 to prevent amorphous or fibrillar aggregation of target proteins was enhanced and correlated with the amount of dissociated species present. We propose that, in vivo, phosphorylation promotes oligomer dissociation, thereby enhancing chaperone activity. Our data support a model in which dimers are the chaperone-active component of Hsp27.
Publisher: American Chemical Society (ACS)
Date: 05-05-2023
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: 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: American Chemical Society (ACS)
Date: 16-12-2019
DOI: 10.1021/ACS.ANALCHEM.9B02068
Abstract: Enrichment strategies are designed for the pretreatment of low-abundance glycans and glycopeptides prior to mass spectrometric (MS) analysis. Here, a tip-based strategy is being reported for the enrichment of glycopeptides and glycans using a piperazine modified polymeric monolithic tip. The tip is fabricated using the free radical polymerization. Fast separation (2 min) is achieved under optimized conditions with 20 cycles per step of loading, incubation, washing, and elution followed by MALDI-MS analysis. A total of 25, 22, and 34 glycopeptides covering all glycosylation sites are enriched by the modified tips from tryptic digests of horse radish peroxidase, chicken avidin, and human immunoglobulin G, respectively. Piperazine exhibits high selectivity 1:400 horse radish peroxidase/bovine serum albumin, sensitivity to 100 attomoles, recovery 89.51%, and batch to batch reproducibility (RSD > 1) in glycopeptides enrichment. Piperazine tips also enrich glycans from ovalbumin and human immunoglobulin G. High selectivity (1:1200, ovalbumin/BSA) and detection limit of 100 attomole is attained for glycans and furthermore 58 glycans are enriched from human serum. Thus, piperazine tips can be used as an enrichment tool for swift, cost-effective routine analysis of biological s les for separation of glycopeptides and glycans.
Publisher: American Chemical Society (ACS)
Date: 09-02-2023
Publisher: Royal Society of Chemistry (RSC)
Date: 2022
DOI: 10.1039/D2FO01305C
Abstract: The magenta lilly pilly ( Syzygium paniculatum ) has been characterised as a high-yielding source of phenolic compounds, particularly anthocyanins, with the berry fruit extracts shown to have broad bioactivity of potential benefit to human health.
Publisher: Public Library of Science (PLoS)
Date: 25-03-2015
Publisher: American Chemical Society (ACS)
Date: 04-12-2017
DOI: 10.1021/ACS.ANALCHEM.7B03328
Abstract: The quaternary structure and dynamics of the human small heat-shock protein Hsp27 are linked to its molecular chaperone function and influenced by post-translational modifications, including phosphorylation. Phosphorylation of Hsp27 promotes oligomer dissociation and can enhance chaperone activity. This study explored the impact of phosphorylation on the quaternary structure and dynamics of Hsp27. Using mutations that mimic phosphorylation, and ion mobility mass spectrometry, we show that successive substitutions result in an increase in the conformational heterogeneity of Hsp27 dimers. In contrast, we did not detect any changes in the structure of an Hsp27 12-mer, representative of larger Hsp27 oligomers. Our data suggest that oligomer dissociation and increased flexibility of the dimer contribute to the enhanced chaperone activity of phosphorylated Hsp27. Thus, post-translational modifications such as phosphorylation play a crucial role in modulating both the tertiary and quaternary structure of Hsp27, which is pivotal to its function as a key component of the proteostasis network in cells. Our data demonstrate the utility of ion mobility mass spectrometry for probing the structure and dynamics of heterogeneous proteins.
Publisher: American Chemical Society (ACS)
Date: 13-11-2020
Publisher: American Chemical Society (ACS)
Date: 18-11-2022
Publisher: Wiley
Date: 06-11-2020
DOI: 10.1111/FEBS.15607
No related grants have been discovered for Blagojce Jovcevski.