ORCID Profile
0000-0003-3284-2176
Current Organisation
University of Melbourne
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Research Topics
In Research Link Australia (RLA), "Research Topics" refer to ANZSRC FOR and SEO codes. These topics are either sourced from ANZSRC FOR and SEO codes listed in researchers' related grants or generated by a large language model (LLM) based on their publications.
Top 5 Research Topics
ANZSRC Field of Research (FoR)
Characterisation of Biological Macromolecules | Medicinal and Biomolecular Chemistry | Cellular Interactions (incl. Adhesion, Matrix, Cell Wall) | Nanomaterials | Nanotechnology | Biologically Active Molecules | Chemical Characterisation of Materials | Medicinal and Biomolecular Chemistry not elsewhere classified | Analytical Biochemistry | Nanobiotechnology |
ANZSRC Socio-Economic Objective (SEO)
Expanding Knowledge in the Chemical Sciences | Expanding Knowledge in the Biological Sciences | Expanding Knowledge in Engineering | Expanding Knowledge in the Medical and Health Sciences
Related Links
Publications
Dye-release assay for investigation of antimicrobial peptide activity in a competitive lipid environment
Publisher: Springer Science and Business Media LLC
Date: 07-06-2014
DOI: 10.1007/S00249-014-0970-0
Abstract: A dye-release method for investigating the effect of a competitive lipid environment on the activity of two membrane-disrupting antimicrobial peptides (AMP), maculatin 1.1 and aurein 1.2, is presented. The results support the general conclusion that AMP have greater affinity for negatively charged membranes, for ex le bacterial membranes, than for the neutral membrane surface found in eukaryotic cells, but only within a competitive lipid environment. Indeed, in a single-model membrane environment, both peptides were more potent against neutral vesicles than against charged vesicles. The approach was also used to investigate the effect of pre-incubating the peptides in a neutral lipid environment then introducing charged lipid vesicles. Maculatin was shown to migrate from the neutral lipid bilayers, where pores had already formed, to the charged membrane bilayers. This result was also observed for charged-to-charged bilayers but, interestingly, not for neutral-to-neutral lipid interfaces. Aurein was able to migrate from either lipid environment, indicating weaker binding to lipid membranes, and a different molecular mechanism for lysis of lipid bilayers. Competitive lipid environments could be used to assess other critical conditions that modulate the activity of membrane peptides or proteins.
Interactions of the antimicrobial peptide maculatin 1.1 and analogues with phospholipid bilayers
Publisher: CSIRO Publishing
Date: 2011
DOI: 10.1071/CH11062
Abstract: The interactions of the antimicrobial peptide, maculatin 1.1 (GLFGVLAKVAAHVVPAIAEHF-NH2) and two analogues, with model phospholipid membranes have been studied using solid-state NMR and circular dichroism spectroscopy. Maculatin 1.1 and the P15G and P15A analogues displayed minimal secondary structure in water, but with zwitterionic dimyristoylphosphatidylcholine (DMPC) vesicles displayed a significant increase in α-helical content. In mixed phospholipid vesicles of DMPC and anionic dimyristoylphosphatidylglycerol (DMPG), each peptide was highly structured with ~80% α-helical content. In DMPC vesicles, the native peptide displayed moderate head group interaction and significant perturbation of the lipid acyl chains. In DMPC/DMPG vesicles, maculatin 1.1 promoted formation of a DMPG-enriched phase and moderately increased disorder towards acyl chain ends of DMPC in the mixed bilayer. Both analogues showed reduced phospholipid head group interactions with DMPC but displayed significant interactions with the mixed lipid system. These effects support the preferential activity of these antimicrobial peptides for bacterial membranes.
19F NMR studies provide insights into lipid membrane interactions of listeriolysin O, a pore forming toxin from Listeria monocytogenes
Publisher: Springer Science and Business Media LLC
Date: 02-05-2018
DOI: 10.1038/S41598-018-24692-6
Abstract: Listeria monocytogenes is a mammalian pathogen that causes gastroenteritis, miscarriages and infections of the central nervous system in immunocompromised in iduals. Its main virulence factor is listeriolysin O (LLO), a pore-forming cholesterol-dependent cytolysin (CDC), which enables bacterial escape from the phagolysosome and contributes to bacterial pathogenicity. Details of cholesterol (Chol) recognition and membrane binding mechanisms by LLO are still not known. Here we used 19 F-NMR spectroscopy in order to assess LLO-Chol interactions in solution and in a Chol-rich membrane environment. LLO has six tryptophan residues located in the region of the molecule that is first in contact with lipid membranes. 19 F-LLO, which contained 5-fluoro-tryptophans, was prepared by using isotopic labelling in an E. coli expression system. Signals in the 19 F-NMR spectrum of 19 F-LLO were unambiguously assigned by using a series of single Trp → Phe point mutations. The results employing various cholesterol preparations in solution indicate that tryptophan residues are not directly involved in Chol binding in solution. However, significant chemical shift changes were observed upon LLO binding to Chol-rich membranes, highlighting the role of tryptophan residues in membrane interactions (W512) and oligomerisation (W189 and W489).
Utilizing magnetic resonance techniques to study membrane interactions of amyloid peptides
Publisher: Portland Press Ltd.
Date: 22-06-2021
DOI: 10.1042/BST20201244
Abstract: Alzheimer's disease (AD) is a common neurodegenerative condition that involves the extracellular accumulation of amyloid plaques predominantly consisting of Aβ peptide aggregates. The amyloid plaques and soluble oligomeric species of Aβ are believed to be the major cause of synaptic dysfunction in AD brain and their cytotoxic mechanisms have been proposed to involve interactions with cell membranes. In this review, we discuss our solid-state nuclear magnetic resonance (ssNMR) studies of Aβ interactions with model membranes.
In-Cell Solid-State NMR Analysis of Membrane Proteins
Publisher: CSIRO Publishing
Date: 15-12-2020
DOI: 10.1071/CH20330
Restriction of lipid motion in membranes triggered by β‐sheet aggregation of the anti‐apoptotic BH4 domain
Publisher: Wiley
Date: 08-01-2008
DOI: 10.1111/J.1742-4658.2007.06222.X
Abstract: The regulative BH4 domain of human Bcl-2 protein exerts its anti-apoptic activity via the mitochondrion. In the present study, we investigated the molecular interactions of this domain with negatively charged liposomes mimicking the outer mitochondrial membrane. To model the overproduction of Bcl-2 found in cancer processes, we studied the impact of elevated concentrations of its regulative BH4 segment on these mitochondrial membranes from the peptide and lipid perspective. Combined solid state (2)H-NMR and differential scanning calorimetry revealed the coexistence of small sized fluid and rigid membrane domains over a large temperature range, which is confirmed by (31)P-NMR at 30 degrees C. The latter are stabilized, in a cholesterol-like manner, by the presence of a BH4 peptide. In the same time scale, the reduction of the headgroup order is seen in the static (14)N and (31)P-NMR spectra when BH4 inserts into the bilayers. Indeed, attenuated total reflection spectroscopy indicated a dominant aggregated beta-sheet secondary structure of BH4 with a 42 degrees tilt relative to the membrane surface. These results are discussed in terms of membrane stabilization versus apoptotic mechanisms at the outer mitochondrial membrane location.
Synthesis of stoichiometrically controlled reactive aluminosilicate and calcium-aluminosilicate powders
Publisher: Elsevier BV
Date: 09-2016
Membrane defects enhance the interaction of antimicrobial peptides, aurein 1.2 versus caerin 1.1
Publisher: Elsevier BV
Date: 08-2013
DOI: 10.1016/J.BBAMEM.2013.03.010
Abstract: The membrane interactions of the antimicrobial peptides aurein 1.2 and caerin 1.1 were observed by (31)P and (2)H solid-state NMR and circular dichroism spectroscopy. Both peptides were relatively unstructured in water. In the presence of dimyristoylphosphatidylcholine (DMPC) and mixed DMPC and dimyristoylphosphatidylglycerol (DMPG) vesicles, both peptides displayed a considerable increase in helical content with the shorter aurein peptide having a higher α-helix content in both lipid systems. In fluid phase DMPC vesicles, the peptides displayed differential interactions: aurein 1.2 interacted primarily with the bilayer surface, while the longer caerin 1.1 was able to penetrate into the bilayer interior. Both peptides displayed a preferential interaction with the DMPG component in DMPC/DMPG bilayers, with aurein 1.2 limited to interaction with the surface and caerin 1.1 able to penetrate into the bilayer and promote formation of a mixture of lipid phases or domains. In gel phase DMPC vesicles, aurein 1.2 disrupted the bilayer apparently through a carpet mechanism, while no additional interaction was seen with caerin 1.1. Although a lamellar bilayer was retained with the mixed DMPC/DMPG vesicles below the phase transition, both caerin 1.1 and aurein 1.2 promoted disruption of the bilayer and formation of an isotropic phase. The peptide interaction was enhanced relative to the fluid phase and was likely driven by co-existence of membrane defects. This study thus demonstrates that the effects of the lipid phase and domains need to be considered when studying membrane interactions of antimicrobial peptides.
Total Chemical Synthesis of an Intra‐A‐Chain Cystathionine Human Insulin Analogue with Enhanced Thermal Stability
Publisher: Wiley
Date: 20-10-2016
The impact of antibacterial peptides on bacterial lipid membranes depends on stage of growth.
Publisher: Royal Society of Chemistry (RSC)
Date: 2021
DOI: 10.1039/D0FD00052C
Abstract: The impact of maculatin 1.1 (Mac1) on the mechanical properties of supported lipid membranes derived from exponential growth phase (EGP) and stationary growth phase (SGP)
Interaction of cationic antimicrobial peptides from Australian frogs with lipid membranes
Publisher: Wiley
Date: 14-03-2018
DOI: 10.1002/PEP2.24061
Enhancing proline-rich antimicrobial peptide action by homodimerization: influence of bifunctional linker
Publisher: Royal Society of Chemistry (RSC)
Date: 2022
DOI: 10.1039/D1SC05662J
Abstract: Homodimerization of a proline-rich antimicrobial peptide via bioconjugation to perfluoroaromatic linkers confers increased antimicrobial, antibiofilm and immunomodulatory activity. The dimers are promising new therapeutic leads against WHO priority multidrug resistant bacteria.
Melittin peptides exhibit different activity on different cells and model membranes
Publisher: Springer Science and Business Media LLC
Date: 09-09-2014
DOI: 10.1007/S00726-014-1833-9
Abstract: Melittin (MLT) is a lytic peptide with a broad spectrum of activity against both eukaryotic and prokaryotic cells. To understand the role of proline and the thiol group of cysteine in the cytolytic activity of MLT, native MLT and cysteine-containing analogs were prepared using solid phase peptide synthesis. The antimicrobial and cytolytic activities of the monomeric and dimeric MLT peptides against different cells and model membranes were investigated. The results indicated that the proline residue was necessary for antimicrobial activity and cytotoxicity and its absence significantly reduced lysis of model membranes and hemolysis. Although lytic activity against model membranes decreased for the MLT dimer, hemolytic activity was increased. The native peptide and the MLT-P14C monomer were mainly unstructured in buffer while the dimer adopted a helical conformation. In the presence of neutral and negatively charged vesicles, the helical content of the three peptides was significantly increased. The lytic activity, therefore, is not correlated to the secondary structure of the peptides and, more particularly, on the propensity to adopt helical conformation.
Prospects for nuclear spin hyperpolarization of molecular samples using nitrogen-vacancy centers in diamond
Publisher: American Physical Society (APS)
Date: 21-01-2021
Orientation and Location of the Cyclotide Kalata B1 in Lipid Bilayers Revealed by Solid-State NMR
Publisher: Elsevier BV
Date: 02-2017
Stoichiometrically controlled C–(A)–S–H/N–A–S–H gel blends via alkali-activation of synthetic precursors
Publisher: Informa UK Limited
Date: 03-08-2015
Proline facilitates membrane insertion of the antimicrobial peptide maculatin 1.1 via surface indentation and subsequent lipid disordering
Publisher: Elsevier BV
Date: 04-2013
Chemical Synthesis and Characterization of an Equinatoxin II(1–85) Analogue
Publisher: MDPI AG
Date: 30-03-2017
Spectroscopic study of L-DOPA and dopamine binding on novel gold nanoparticles towards more efficient drug-delivery system for Parkinson’s disease
Publisher: Elsevier BV
Date: 03-2022
DOI: 10.1016/J.SAA.2021.120707
Abstract: Nano-drug delivery systems may potentially overcome current challenges in the treatment of Parkinson's disease (PD) by enabling targeted delivery and more efficient blood-brain penetration ability. This study investigates novel gold nanoparticles (AuNPs) to be used as delivery systems for L-DOPA and dopamine by considering their binding capabilities in the presence and absence of a model protein, bovine serum albumin (BSA). Four different AuNPs were prepared by surface functionalization with polyethylene glycol (PEG), 1-adamantylamine (Ad), 1-adamantylglycine (AdGly), and peptidoglycan monomer (PGM). Fluorescence and UV-Vis measurements demonstrated the strongest binding affinity and L-DOPA/dopamine loading efficiency for PGM-functionalized AuNPs with negligible impact of the serum protein presence. Thermodynamic analysis revealed a spontaneous binding process between L-DOPA or dopamine and AuNPs that predominantly occurred through van der Waals interactions/hydrogen bonds or electrostatic interactions. These results represent PGM-functionalized AuNPs as the most efficient at L-DOPA and dopamine binding with a potential to become a drug-delivery system for neurodegenerative diseases. Detailed investigation of L-DOPA/dopamine interactions with different AuNPs was described here for the first time. Moreover, this study highlights a cost- and time-effective methodology for evaluating drug binding to nanomaterials.
Selenium Nanoparticles as Potential Drug-Delivery Systems for the Treatment of Parkinson’s Disease
Publisher: American Chemical Society (ACS)
Date: 20-09-2023
The Prototypic Cyclotide Kalata B1 Has a Unique Mechanism of Entering Cells
Publisher: Elsevier BV
Date: 08-2015
DOI: 10.1016/J.CHEMBIOL.2015.07.012
Abstract: Cyclotides combine the stability of disulfide-rich peptides with the intracellular accessibility of cell-penetrating peptides, giving them outstanding potential as drug scaffolds with an ability to inhibit intracellular protein-protein interactions. To realize and optimize the application of cyclotides as a drug framework and delivery system, we studied the ability of the prototypic cyclotide, kalata B1, to enter mammalian cells. We show that kalata B1 can enter cells via both endocytosis and direct membrane translocation. Both pathways are initiated by targeting phosphatidylethanolamine phospholipids at the cell surface and inducing membrane curvature. This unusual approach to initiate internalization might be harnessed to deliver drugs into cells and, in particular, cancer cells, which present a higher proportion of surface-exposed phosphatidylethanolamine phospholipids. Our findings highlight the potential of these peptides as drug leads for the modulation of traditionally "undruggable" targets, such as intracellular protein-protein interactions.
How is protein aggregation in amyloidogenic diseases modulated by biological membranes?
Publisher: Springer Science and Business Media LLC
Date: 21-11-2007
DOI: 10.1007/S00249-007-0237-0
Abstract: The fate of proteins with amyloidogenic properties depends critically on their immediate biochemical environment. However, the role of biological interfaces such as membrane surfaces, as promoters of pathological aggregation of amyloidogenic proteins, is rarely studied and only established for the amyloid-beta protein (A beta) involved in Alzheimer's disease, and alpha-synuclein in Parkinsonism. The occurrence of binding and misfolding of these proteins on membrane surfaces, is poorly understood, not at least due to the two-dimensional character of this event. Clearly, the nature of the folding pathway for A beta protein adsorbed upon two-dimensional aggregation templates, must be fundamentally different from the three-dimensional situation in solution. Here, we summarize the current research and focus on the function of membrane interfaces as aggregation templates for amyloidogenic proteins (and even prionic ones). One major aspect will be the relationship between membrane properties and protein association and the consequences for amyloidogenic products. The other focus will be on a general understanding of protein folding pathways on two-dimensional templates on a molecular level. Finally, we will demonstrate the potential importance of membrane-mediated aggregation for non- hiphatic soluble amyloidogenic proteins, by using the SOD1 protein involved in the amyotrophic lateral sclerosis syndrome.
Phase evolution of Na2O–Al2O3–SiO2–H2O gels in synthetic aluminosilicate binders
Publisher: Royal Society of Chemistry (RSC)
Date: 2016
DOI: 10.1039/C5DT04878H
Abstract: Nanostructural evolution of Na 2 O–Al 2 O 3 –SiO 2 –H 2 O gels in synthetic aluminosilicate binders investigated by solid state 29 Si, 27 Al and 23 Na MAS NMR spectroscopy.
Expression and purification of the native C‐amidated antimicrobial peptide maculatin 1.1
Publisher: Wiley
Date: 11-04-2021
DOI: 10.1002/PSC.3330
Abstract: Maculatin 1.1 (Mac1) is an antimicrobial peptide (AMP) from an Australian tree frog and exhibits low micromolar activity against Gram‐positive bacteria. The antimicrobial properties of Mac1 are linked to its disruption of bacterial lipid membranes, which has been studied extensively by in vitro nuclear magnetic resonance (NMR) spectroscopy and biophysical approaches. Although in vivo NMR has recently proven effective in probing peptide‐lipid interplay in live bacterial cells, direct structural characterisation of AMPs has been prohibited by low sensitivity and overwhelming background noise. To overcome this issue, we report a recombinant expression protocol to produce isotopically enriched Mac1. We utilized a double‐fusion construct to alleviate toxicity against the Escherichia coli host and generate the native N‐free and C‐amidated termini Mac1 peptide. The SUMO and intein tags allowed native N‐terminus and C‐terminal amidation, respectively, to be achieved in a one‐pot reaction. The protocol yielded 0.1 mg/L of native, uniformly 15 N‐labelled, Mac1, which possessed identical structure and activity to peptide obtained by solid‐phase peptide synthesis.
The Location of the Antimicrobial Peptide Maculatin 1.1 in Model Bacterial Membranes
Publisher: Frontiers Media SA
Date: 07-07-2020
Aggregation kinetics in the presence of brain lipids of Aβ(1–40) cleaved from a soluble fusion protein
Publisher: Elsevier BV
Date: 09-2018
DOI: 10.1016/J.BBAMEM.2018.03.005
Abstract: The cleavage of the amyloid precursor protein by β- and γ-secretases is a key event in Alzheimer's disease. A fusion protein was constructed to investigate the cleavage rate and aggregation kinetics of amyloid-beta (1-40) (Aβ(1-40)) peptides. The peptide was expressed with a Small Ubiquitin-Like Modifier (SUMO) on the N-terminus and cleaved by a SUMO protease Ulp1. The time course of the cleavage reaction was monitored by SDS-PAGE gel with 100:1 or 1000:1 SUMO-Aβ(1-40) to Ulp1 molar ratio and in the presence of brain total lipid extract unilamellar vesicles. Similarly, the aggregation of Aβ(1-40) peptides upon cleavage was monitored by thioflavin T fluorescence assays and by circular dichroism. The cleavage reaction was modulated by the concentration of Ulp1, with fast release of Aβ(1-40) peptides producing shorter lag time before fibril formation, but with similar elongation rate. The presence of lipids significantly reduced the cleavage completion at 1000:1, but reduced the lag time before fibril formation, while at 100:1 similar cleavage and aggregation kinetics were observed compared to the lipid-free condition. Overall, the results showed that the fusion protein SUMO-Aβ(1-40) is a means to study the cleavage and aggregation of amyloid peptides and that the presence of lipids and the fast release rate accelerated the aggregation of Aβ(1-40) peptides.
C-terminus amidation influences biological activity and membrane interaction of maculatin 1.1
Publisher: Springer Science and Business Media LLC
Date: 23-04-2021
DOI: 10.1007/S00726-021-02983-Z
Abstract: Cationic antimicrobial peptides have been investigated for their potential use in combating infections by targeting the cell membrane of microbes. Their unique chemical structure has been investigated to understand their mode of action and optimize their dose-response by rationale design. One common feature among cationic AMPs is an amidated C-terminus that provides greater stability against in vivo degradation. This chemical modification also likely modulates the interaction with the cell membrane of bacteria yet few studies have been performed comparing the effect of the capping groups. We used maculatin 1.1 (Mac1) to assess the role of the capping groups in modulating the peptide bacterial efficiency, stability and interactions with lipid membranes. Circular dichroism results showed that C-terminus amidation maintains the structural stability of the peptide (α-helix) in contact with micelles. Dye leakage experiments revealed that amidation of the C-terminus resulted in higher membrane disruptive ability while bacteria and cell viability assays revealed that the amidated form displayed higher antibacterial ability and cytotoxicity compared to the acidic form of Mac1. Furthermore,
TOAC spin-labeled peptides tailored for DNP-NMR studies in lipid membrane environments
Publisher: Elsevier BV
Date: 10-2021
How Membrane-Active Peptides Get into Lipid Membranes
Publisher: American Chemical Society (ACS)
Date: 17-05-2016
DOI: 10.1021/ACS.ACCOUNTS.6B00074
Abstract: The structure-function relationship for a family of antimicrobial peptides (AMPs) from the skin of Australian tree frogs is discussed and compared with that of peptide toxins from bee and Australian scorpion venoms. Although these membrane-active peptides induce a similar cellular fate by disrupting the lipid bilayer integrity, their lytic activity is achieved via different modes of action, which are investigated in relation to amino acid sequence, secondary structure, and membrane lipid composition. In order to better understand what structural features govern the interaction between peptides and lipid membranes, cell-penetrating peptides (CPPs), which translocate through the membrane without compromising its integrity, are also discussed. AMPs possess membrane lytic activities that are naturally designed to target the cellular membrane of pathogens or competitors. They are extremely erse in amino acid composition and often show specificity against a particular strain of microbe. Since our antibiotic arsenal is declining precariously in the face of the rise in multiantibiotic resistance, AMPs increasingly are seen as a promising alternative. In an effort to understand their molecular mechanism, biophysical studies of a myriad of AMPs have been reported, yet no unifying mechanism has emerged, rendering difficult the rational design of drug leads. Similarly, a wide variety of cytotoxic peptides are found in venoms, the best known being melittin, yet again, predicting their activity based on a particular amino acid composition or secondary structure remains elusive. A common feature of these membrane-active peptides is their preference for the lipid environment. Indeed, they are mainly unstructured in solution and, in the presence of lipid membranes, quickly adsorb onto the surface, change their secondary structure, eventually insert into the hydrophobic core of the membrane bilayer, and finally disrupt the bilayer integrity. These steps define the molecular mechanism by which these membrane-active peptides lyse membranes. The last class of membrane-active peptides discussed are the CPPs, which translocate across the lipid bilayer without inducing severe disruption and have potential as drug vehicles. CPPs are typically highly charged and can show antimicrobial activity by targeting an intracellular target rather than via a direct membrane lytic mechanism. A critical aspect in the structure-function relationship of membrane-active peptides is their specific activity relative to the lipid membrane composition of the cell target. Cell membranes have a wide ersity of lipids, and those of eukaryotic and prokaryotic species differ greatly in composition and structure. The activity of AMPs from Australian tree frogs, toxins, and CPPs has been investigated within various lipid systems to assess whether a relationship between peptide and membrane composition could be identified. NMR spectroscopy techniques are being used to gain atomistic details of how these membrane-active peptides interact with model membranes and cells, and in particular, competitive assays demonstrate the difference between affinity and activity for a specific lipid environment. Overall, the interactions between these relatively small sized peptides and various lipid bilayers give insight into how these peptides function at the membrane interface.
Structural Disruptions of the Outer Membranes of Gram-Negative Bacteria by Rationally Designed Amphiphilic Antimicrobial Peptides
Publisher: American Chemical Society (ACS)
Date: 02-04-2021
Interaction of N-terminal peptide analogues of the Na+,K+-ATPase with membranes
Publisher: Elsevier BV
Date: 06-2018
DOI: 10.1016/J.BBAMEM.2018.03.002
Abstract: The Na
Probing Ionic Liquid Electrolyte Structure via the Glassy State by Dynamic Nuclear Polarization NMR Spectroscopy
Publisher: American Chemical Society (ACS)
Date: 13-02-2018
DOI: 10.1021/ACS.JPCLETT.8B00022
Abstract: Dynamic nuclear polarization (DNP)-enhanced solid-state NMR spectroscopy has been used to study an ionic liquid salt solution (N-methyl-N-propyl-pyrrolidinium bis(fluorosulfonyl)imide, C
Bacteria May Cope Differently from Similar Membrane Damage Caused by the Australian Tree Frog Antimicrobial Peptide Maculatin 1.1
Publisher: Elsevier BV
Date: 08-2015
Listeriolysin O Binding Affects Cholesterol and Phospholipid Acyl Chain Dynamics in Fluid Cholesterol-Rich Bilayers.
Publisher: Wiley
Date: 28-08-2018
Abstract: Listeriolysin O (LLO) is a pore-forming toxin that enables survival and cell-to-cell spread of foodborne bacterial pathogen Listeria monocytogenes, which is responsible for the life-threatening disease, listeriosis. LLO-membrane interactions are crucial for pathogenicity of Listeria, but remained unexplained in detail at the molecular level. Here we addressed them by means of
DNP NMR spectroscopy reveals new structures, residues and interactions in wild spider silks
Publisher: Royal Society of Chemistry (RSC)
Date: 2019
DOI: 10.1039/C9CC01045A
Abstract: DNP solid state NMR spectroscopy allows non-targeted analysis of wild spider silk in unprecedented detail at natural abundance, revealing hitherto unreported features across several species.
Examination of alkali-activated material nanostructure during thermal treatment
Publisher: Springer Science and Business Media LLC
Date: 05-04-2018
A New Turn in Peptide-Based Imaging Agents: Foldamers Afford Improved Theranostics Targeting Cholecystokinin-2 Receptor-Positive Cancer
Publisher: American Chemical Society (ACS)
Date: 07-04-2021
Total Chemical Synthesis of an Intra‐A‐Chain Cystathionine Human Insulin Analogue with Enhanced Thermal Stability
Publisher: Wiley
Date: 20-10-2016
Abstract: Despite recent advances in the treatment of diabetes mellitus, storage of insulin formulations at 4 °C is still necessary to minimize chemical degradation. This is problematic in tropical regions where reliable refrigeration is not ubiquitous. Some degradation byproducts are caused by disulfide shuffling of cystine that leads to covalently bonded oligomers. Consequently we examined the utility of the non‐reducible cystine isostere, cystathionine, within the A‐chain. Reported herein is an efficient method for forming this mimic using simple monomeric building blocks. The intra‐A‐chain cystathionine insulin analogue was obtained in good overall yield, chemically characterized and demonstrated to possess native binding affinity for the insulin receptor isoform B. It was also shown to possess significantly enhanced thermal stability indicating potential application to next‐generation insulin analogues.
Progression of NMR studies of membrane-active peptides from lipid bilayers to live cells
Publisher: Elsevier BV
Date: 04-2015
DOI: 10.1016/J.JMR.2014.11.016
Abstract: Understanding the structure of membrane-active peptides faces many challenges associated with the development of appropriate model membrane systems as the peptide structure depends strongly on the lipid environment. This perspective provides a brief overview of the approach taken to study antimicrobial and amyloid peptides in phospholipid bilayers using oriented bilayers and magic angle spinning techniques. In particular, Boltzmann statistics REDOR and maximum entropy analysis of spinning side bands are used to analyse systems where multiple states of peptide or lipid molecules may co-exist. We propose that in future, rather than model membranes, structural studies in whole cells are feasible.
Membrane Interacting Peptides: From Killers to Helpers
Publisher: Bentham Science Publishers Ltd.
Date: 11-2012
DOI: 10.2174/138920312804142138
Abstract: Membrane interacting peptides are reviewed in terms of structure and mode of action on lipid membranes. Helical, β-stranded, peptides containing both helices and strands, cyclic, lipopeptides and short linear peptides are seen to considerably modulate membrane function. Among peptides that lead to membrane alteration or permeation, antimicrobial peptides play an important role and some of them may be foreseen as potential new antibiotics. Alternatively, peptides that do not destroy the membrane are also very important in modulating the structure and dynamics of the lipid bilayer and play important roles in membrane protein functions. Peptide lipid complexes are shown to be very variable in structure and dynamics: "carpet", "barrel stave", toroid and disordered pores, electrostatic wedge and molecular electroporation models are discussed. Their assembly is reviewed in terms of electric, hipathic and dynamic properties of both lipids and peptides.
Structural evolution of synthetic alkali-activated CaO-MgO-Na 2 O-Al 2 O 3 -SiO 2 materials is influenced by Mg content
Publisher: Elsevier BV
Date: 09-2017
Phase evolution of C-(N)-A-S-H/N-A-S-H gel blends investigated via alkali-activation of synthetic calcium aluminosilicate precursors
Publisher: Elsevier BV
Date: 11-2016
Effect of phosphatidylcholine bilayer thickness and molecular order on the binding of the antimicrobial peptide maculatin 1.1
Publisher: Elsevier BV
Date: 02-2018
DOI: 10.1016/J.BBAMEM.2017.10.007
Abstract: Antimicrobial peptides (AMPs) interact directly with bacterial membrane lipids. Thus, changes in the lipid composition of bacterial membranes can have profound effects on the activity of AMPs. In order to understand the effect of bilayer thickness and molecular order on the activity of AMPs, the interaction of maculatin 1.1 (Mac1.1) with phosphatidylcholine (PC) model membranes composed of different monounsaturated acyl chain lengths between 14 and 22 carbons was characterised by dual polarisation interferometry (DPI) and 31P and 1H solid-state NMR techniques. The thickness and bilayer order of each PC bilayer showed a linear dependence on the acyl chain length. The binding of Mac1.1 exhibited a biphasic dependency between the amount of bound Mac1.1 and bilayer thickness, whereby the mass of bound peptide increased from C14 to C16 and then decreased from C16 to C22. Significant perturbation of 31P chemical shift anisotropy (CSA) values was only observed for DOPC (C18) and DEPC (C22), respectively. In the case of DEPC, the greater range in CSA indicated different headgroup conformations or environments in the presence of Mac1.1. Overall, the results indicated that there is a significant change in the bilayer order upon binding of Mac1.1 and this change occurred in a co-operative manner at higher concentrations of Mac1.1 with increasing bilayer thickness and order. Overall, an optimum bilayer thickness and lipid order may be required for effective membrane perturbation by Mac1.1 and increasing the bilayer thickness and order may counteract the activity of Mac1.1 and play a role in antimicrobial resistance to AMPs.
Phote-HrTH (Phormia terraenovae Hypertrehalosaemic Hormone), the Metabolic Hormone of the Fruit Fly: Solution Structure and Receptor Binding Model
Publisher: CSIRO Publishing
Date: 2020
DOI: 10.1071/CH19461
Abstract: Fruit flies are a widely distributed pest insect that pose a significant threat to food security. Flight is essential for the dispersal of the adult flies to find new food sources and ideal breeding spots. The supply of metabolic fuel to power the flight muscles of insects is regulated by adipokinetic hormones (AKHs). The fruit fly, Drosophila melanogaster, has the same AKH that is present in the blowfly, Phormia terraenovae this AKH has the code-name Phote-HrTH. Binding of the AKH to the extra-cellular binding site of a G protein-coupled receptor causes its activation. In this paper, the structure of Phote-HrTH in sodium dodecyl sulfate (SDS) micelle solution was determined using NMR restrained molecular dynamics. The peptide was found to bind to the micelle and be fairly rigid, with an S2 order parameter of 0.96. The translated protein sequence of the AKH receptor from the fruit fly, D. melanogaster, Drome-AKHR, was used to construct two models of the receptor. It is proposed that these two models represent the active and inactive state of the receptor. The model based on the crystal structure of the β-2 adrenergic receptor was found to bind Phote-HrTH with a binding constant of −102kJmol−1, while the other model, based on the crystal structure of rhodopsin, did not bind the peptide. Under molecular dynamic simulation, in a palmitoyloleoylphosphatidylcholine (POPC) membrane, the receptor complex changed from an inactive to an active state. The identification and characterisation of the ligand binding site of Drome-AKHR provide novel information of ligand–receptor interaction, which could lead to the development of species-specific control substances to use discriminately against the fruit fly.
Elucidating the bactericidal mechanism of action of the linear antimicrobial tetrapeptide BRBR-NH2
Publisher: Elsevier BV
Date: 08-2018
DOI: 10.1016/J.BBAMEM.2018.05.004
Abstract: Linear antimicrobial peptides, with their rapid bactericidal mode of action, are well-suited for development as topical antibacterial drugs. We recently designed a synthetic linear 4-residue peptide, BRBR-NH
Magic‐angle phosphorus NMR of functional mitochondria:in situmonitoring of lipid response under apoptotic‐like stress
Publisher: Wiley
Date: 24-04-2009
DOI: 10.1096/FJ.09-134114
Abstract: Using a noninvasive, solid-state magic-angle spinning nuclear magnetic resonance (MAS NMR) approach, we track ex vivo the behavior of in idual membrane components in isolated, active mitochondria (model system: potato tubers) during physiological processes. The in idual phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cardiolipin (CL) membrane constituents were identified as distinct lines by applying MAS (31)P NMR on extracted lipid membranes. However, the CL NMR signal appeared to be very broad in functional mitochondria, indicating a tight complex formation with membrane protein. Calcium stress induced severe membrane degradation without recovery of a single CL NMR resonance. This suggests that calcium overload destroys the outer mitochondrial membrane and does not modify strongly the CL protein complexes in the inner membrane a conclusion confirmed by respiratory controls. Conversely, mitochondrial membrane disruption on time degradation or mechanical stress generates clearly visible identical CL NMR signals, similar to those observed in rehydrated lipid extracts. Similarly, noninvasive based NMR tracking of lipids in response to erse physiological stimuli can easily be used for other organelles and whole living cells.
Multi-Omic Analysis to Characterize Metabolic Adaptation of the E. coli Lipidome in Response to Environmental Stress
Publisher: MDPI AG
Date: 11-02-2022
Abstract: As an adaptive survival response to exogenous stress, bacteria undergo dynamic remodelling of their lipid metabolism pathways to alter the composition of their cellular membranes. Here, using Escherichia coli as a well characterised model system, we report the development and application of a ‘multi-omics’ strategy for comprehensive quantitative analysis of the temporal changes in the lipidome and proteome profiles that occur under exponential growth phase versus stationary growth phase conditions i.e., nutrient depletion stress. Lipidome analysis performed using ‘shotgun’ direct infusion-based ultra-high resolution accurate mass spectrometry revealed a quantitative decrease in total lipid content under stationary growth phase conditions, along with a significant increase in the mol% composition of total cardiolipin, and an increase in ‘odd-numbered’ acyl-chain length containing glycerophospholipids. The inclusion of field asymmetry ion mobility spectrometry was shown to enable the enrichment and improved depth of coverage of low-abundance cardiolipins, while ultraviolet photodissociation-tandem mass spectrometry facilitated more complete lipid structural characterisation compared with conventional collision-induced dissociation, including unambiguous assignment of the odd-numbered acyl-chains as containing cyclopropyl modifications. Proteome analysis using data-dependent acquisition nano-liquid chromatography mass spectrometry and tandem mass spectrometry analysis identified 83% of the predicted E. coli lipid metabolism enzymes, which enabled the temporal dependence associated with the expression of key enzymes responsible for the observed adaptive lipid metabolism to be determined, including those involved in phospholipid metabolism (e.g., ClsB and Cfa), fatty acid synthesis (e.g., FabH) and degradation (e.g., FadA/B,D,E,I,J and M), and proteins involved in the oxidative stress response resulting from the generation of reactive oxygen species during β-oxidation or lipid degradation.
NMR Chemical Shift and Methylation of 4-Nitroimidazole: Experiment and Theory
Publisher: CSIRO Publishing
Date: 2021
DOI: 10.1071/CH20199
Abstract: Nitroimidazoles and derivatives are a class of active pharmaceutical ingredients (APIs) first introduced sixty years ago. As anti-infection agents, the structure–activity relationships of nitroimidazole compounds have been particularly difficult to study due to their low reduction potentials and unique electronic structures. In this study, we combine dynamic nuclear polarization (DNP)-enhanced solid-state (100K), solid-state (298K), and 1H-13C heteronuclear single quantum coherence (HSQC) solution-state NMR techniques (303K) with density functional theory (DFT) to study the 1H, 13C, and 15N chemical shifts of 4-nitroimidazole (4-NI) and 1-methyl-4-nitroimidazole (CH3-4NI). The 4-NI chemical shifts were observed at 119.4, 136.4, and 144.7ppm for 13C, and at 181.5, 237.4, and 363.0ppm for 15N. The measurements revealed that methylation (deprotonation) of the amino nitrogen N(1) of 4-NI had less effect (Δδ=−4.8ppm) on the N(1) chemical shift but was compensated by shielding of the N(3) (Δδ=11.6ppm) in CH3-4NI. The calculated chemical shifts using DFT for 4-NI and CH3-4NI agreed well with the experimental values (within 2%) for the imidazole carbons. However, larger discrepancies (up to 13%) were observed between the calculated and measured 15N NMR chemical shifts for the imidazole nitrogen atoms of both molecules, which indicate that effects such as imidazole ring resonant structures and molecular dynamics may also contribute to the nitrogen chemical environment.
Membrane Insertion of a Dinuclear Polypyridylruthenium(II) Complex Revealed by Solid-State NMR and Molecular Dynamics Simulation: Implications for Selective Antibacterial Activity
Publisher: American Chemical Society (ACS)
Date: 09-11-2016
DOI: 10.1021/JACS.6B09996
Abstract: Dinuclear polypyridylruthenium(II) complexes bridged by a flexible methylene linker have received considerable interest as potential antibacterial agents. Their potency and uptake into bacterial cells is directly modulated by the length of the bridging linker, which has implicated membrane interactions as an essential feature of their mechanism of action. In this work, a combination of molecular dynamics (MD) simulations and solid-state NMR was used to present an atomistic model of a polypyridylruthenium(II) complex bound and incorporated into a bacterial membrane model. The results of
How does the Bax-α1 targeting sequence interact with mitochondrial membranes? The role of cardiolipin
Publisher: Elsevier BV
Date: 03-2009
DOI: 10.1016/J.BBAMEM.2008.12.014
Abstract: A key event in programmed cell death is the translocation of the apoptotic Bax protein from the cytosol towards mitochondria. The first helix localized at the N-terminus of Bax (Bax-alpha1) can act here as an addressing sequence, which directs activated Bax towards the mitochondrial surface. Solid state NMR (nuclear magnetic resonance), CD (circular dichroism) and ATR (attenuated total reflection) spectroscopy were used to elucidate this recognition process of a mitochondrial membrane system by Bax-alpha1. Two potential target membranes were studied, with the outer mitochondrial membrane (OM) mimicked by neutral phospholipids, while mitochondrial contact sites (CS) contained additional anionic cardiolipin. (1)H and (31)P magic angle spinning (MAS) NMR revealed Bax-alpha1 induced pronounced perturbations in the lipid headgroup region only in presence of cardiolipin. Bax-alpha1 could not insert into CS membranes but at elevated concentrations it inserted into the hydrophobic core of cardiolipin-free OM vesicles, thereby adopting beta-sheet-like features, as confirmed by ATR. CD studies revealed, that the cardiolipin mediated electrostatic locking of Bax-alpha1 at the CS membrane surface promotes conformational changes into an alpha-helical state a process which seems to be necessary to induce further conformational transition events in activated Bax which finally causes irreversible membrane permeabilization during the mitochondrial apoptosis.
Disordered Proteins: Biological Membranes as Two-Dimensional Aggregation Matrices
Publisher: Springer Science and Business Media LLC
Date: 31-10-2008
DOI: 10.1007/S12013-008-9033-4
Abstract: Aberrant folded proteins and peptides are hallmarks of amyloidogenic diseases. However, the molecular processes that cause these proteins to adopt non-native structures in vivo and become cytotoxic are still largely unknown, despite intense efforts to establish a general molecular description of their behavior. Clearly, the fate of these proteins is ultimately linked to their immediate biochemical environment in vivo. In this review, we focus on the role of biological membranes, reactive interfaces that not only affect the conformational stability of amyloidogenic proteins, but also their aggregation rates and, probably, their toxicity. We first provide an overview of recent work, starting with findings regarding the hiphatic amyloid-beta protein (Abeta), which give evidence that membranes can directly promote aggregation, and that the effectiveness in this process can be related to the presence of specific neuronal ganglioside lipids. In addition, we discuss the implications of recent research (medin as an detailed ex le) regarding putative roles of membranes in the misfolding behavior of soluble, non- hiphatic proteins, which are attracting increasing interest. The potential role of membranes in exerting the toxic action of misfolded proteins will also be highlighted in a molecular context. In this review, we discuss novel NMR-based approaches for exploring membrane-protein interactions, and findings obtained using them, which we use to develop a molecular concept to describe membrane-mediated protein misfolding as a quasi-two-dimensional process rather than a three-dimensional event in a biochemical environment. The aim of the review is to provide researchers with a general understanding of the involvement of membranes in folding/misfolding processes in vivo, which might be quite universal and important for future research concerning amyloidogenic and misfolding proteins, and possible ways to prevent their toxic actions.
NMR spectroscopy of lipidic cubic phases
Publisher: Springer Science and Business Media LLC
Date: 11-11-2021
Structural Biology, ASB/ABA meeting, Melbourne
Publisher: Springer Science and Business Media LLC
Date: 29-04-2019
Structure, Function, and Biosynthetic Origin of Octapeptin Antibiotics Active against Extensively Drug-Resistant Gram-Negative Bacteria
Publisher: Elsevier BV
Date: 04-2018
One pathogen two stones: are Australian tree frog antimicrobial peptides synergistic against human pathogens?
Publisher: Springer Science and Business Media LLC
Date: 06-05-2017
DOI: 10.1007/S00249-017-1215-9
Abstract: Antimicrobial peptides (AMPs) may act by targeting the lipid membranes and disrupting the bilayer structure. In this study, three AMPs from the skin of Australian tree frogs, aurein 1.2, maculatin 1.1 and caerin 1.1, were investigated against Gram-negative Escherichia coli, Gram-positive Staphylococcus aureus, and vesicles that mimic their lipid compositions. Furthermore, equimolar mixtures of the peptides were tested to identify any synergistic interactions in antimicrobial activity. Minimum inhibition concentration and minimum bactericidal concentration assays showed significant activity against S. aureus but not against E. coli. Aurein was the least active while maculatin was the most active peptide and some synergistic effects were observed against S. aureus. Circular dichroism experiments showed that, in the presence of phospholipid vesicles, the peptides transitioned from an unstructured to a predominantly helical conformation (>50%), with greater helicity for POPG/TOCL compared to POPE/POPG vesicles. The helical content, however, was less in the presence of live E. coli and S. aureus, 25 and 5%, respectively. Equimolar concentrations of the peptides did not appear to form greater supramolecular structures. Dye release assays showed that aurein required greater concentration than caerin and maculatin to disrupt the lipid bilayers, and mixtures of the peptides did not cooperate to enhance their lytic activity. Overall, aurein, maculatin, and caerin showed moderate synergy in antimicrobial activity against S. aureus without becoming more structured or enhancement of their membrane-disrupting activity in phospholipid vesicles.
Star Polymers Reduce Islet Amyloid Polypeptide Toxicity via Accelerated Amyloid Aggregation
Publisher: American Chemical Society (ACS)
Date: 31-10-2017
The antimicrobial peptide maculatin self assembles in parallel to form a pore in phospholipid bilayers
Publisher: Elsevier BV
Date: 05-2020
DOI: 10.1016/J.BBAMEM.2020.183204
Abstract: Little is known experimentally about the detailed orientation of membrane-bound maculatin 1.1 (Mac1), an antimicrobial peptide from the skin secretions of Australian tree frogs. In this work multiple
Hypercrosslinked additives for ageless gas-separation membranes
Publisher: Wiley
Date: 08-01-2016
Abstract: The loss of internal pores, a process known as physical aging, inhibits the long-term use of the most promising gas-separation polymers. Previously we reported that a porous aromatic framework (PAF-1) could form a remarkable nanocomposite with gas-separation polymers to stop aging. However, PAF-1 synthesis is very onerous both from a reagent and reaction-condition perspective, making it difficult to scale-up. We now reveal a highly dispersible and scalable additive based on α,α'-dichloro-p-xylene (p-DCX), that inhibits aging more effectively, and crucially almost doubles gas-transport selectivity. These synergistic effects are related to the intimately mixed nanocomposite that is formed though the high dispersibility of p-DCX in the gas-separation polymer. This reduces particle-size effects and the internal free volume is almost unchanged over time. This study shows this inexpensive and scalable polymer additive delivers exceptional gas-transport performance and selectivity.
Membrane interactions of proline-rich antimicrobial peptide, Chex1-Arg20, multimers
Publisher: Elsevier BV
Date: 06-2016
DOI: 10.1016/J.BBAMEM.2016.02.035
Abstract: The increasing prevalence of antibiotic-resistant pathogens requires the development of new antibiotics. Proline-rich antimicrobial peptides (PrAMPs), including native apidaecins, Bac7, and oncocins or designed A3APO, show multi-modal actions against pathogens together with immunostimulatory activities. The interactions of the designed PrAMP, Chex1-Arg20, and its dimeric and tetrameric oligomers with different model membranes were investigated by circular dichroism spectroscopy, dynamic light scattering, zeta potential, differential scanning calorimetry, and dye leakage. Chex1-Arg20 oligomers showed stronger affinity and preferential binding to negatively charged phospholipid bilayers and led to lipid aggregation and neutralization. Fluorescence microscopy of negatively charged giant unilamellar vesicles with AlexFluor-647-labeled Chex1-Arg20 dimers and tetramers displayed aggregation at a peptide/lipid low ratio of 1:200 and at higher peptide concentrations (1:100/1:50) for Chex1-Arg20 monomer. Such interactions, aggregation, and neutralization of PrAMP oligomers additionally showed the importance of interactions of PrAMPs with negatively charged membranes.
Glycine Substitution Reduces Antimicrobial Activity and Helical Stretch of diPGLa-H in Lipid Micelles
Publisher: American Chemical Society (ACS)
Date: 28-04-2017
Abstract: With the rise in antibiotic resistance, antimicrobial peptides (AMPs) show promise for therapeutic development, but higher specificity is required. PGLa-H is a naturally occurring decapeptide, reported to have moderate antibacterial activity and low hemolytic activity, with its sequence being identical to that of the C-terminal fragment of highly selective AMP, PGLa. DiPGLa-H, a sequential tandem repeat of PGLa-H, and Kiadin, an analogue with a Val to Gly substitution at position 15, display improved in vitro bactericidal activity against both Gram-negative and Gram-positive pathogens, with generally low toxicity for human cells. Despite Gly being a more flexible residue, NMR structural studies showed little difference in structure and dynamics between the two peptides for the first 14 residues, with somewhat greater flexibility in the C-terminus of Kiadin resulting in a tighter structure of the peptide in the presence of sodium dodecyl sulfate micelles. AMPs found in organisms often exhibit minimal amino acid mutations, and such small differences in peptide conformation may be utilized to design more selective AMPs.
Copolyampholytes Produced from RAFT Polymerization of Protic Ionic Liquids
Publisher: American Chemical Society (ACS)
Date: 10-11-2017
Proline-15 creates an amphipathic wedge in maculatin 1.1 peptides that drives lipid membrane disruption
Publisher: Elsevier BV
Date: 10-2015
DOI: 10.1016/J.BBAMEM.2015.06.013
Abstract: The membrane interaction of peptides derived from maculatin 1.1 and caerin 1.1, with the sequence motif of N and C termini of maculatin 1.1, was compared in order to understand the role of these common sequence motifs, which encompass critical proline residues, on peptide secondary structure and on membrane binding and disruption in zwitterionic and anionic membranes. The peptides incorporated a single substitution with lysine or deletion of the central region to mimic the length of the antimicrobial peptides, citropin 1.1 and aurein 1.2. The impact of these changes in the sequence, length and physicochemical properties, on lytic activity and structure was assessed by dye-release from lipid vesicles and the change in the bilayer order as a function of membrane-bound peptide mass. All peptides adopted similar degrees of helical structure in both membrane systems. In addition, all peptide analogues were less active than either maculatin 1.1 or caerin 1.1 in dye release assays. The membrane binding was analyzed by dual polarization interferometry and the results showed that membrane binding was significantly affected by changes in the hydrophobic environment of Pro-15. Moreover, changes in the relative distribution of charge and hydrophobicity flanking Pro-15 also caused significant changes to the membrane order. Overall, the proline residue plays an important role in inducing a peptide structure that enhances the activity of these antimicrobial peptides.
A multifunctional surfactant catalyst inspired by hydrolases
Publisher: American Association for the Advancement of Science (AAAS)
Date: 03-04-2020
Abstract: Enzyme-inspired catalysis using self-assembled hiphiles mimic and localize multiple chemical units common to hydrolases.
In Situ Monitoring of Bacteria under Antimicrobial Stress Using 31P Solid-State NMR
Publisher: MDPI AG
Date: 06-01-2019
DOI: 10.3390/IJMS20010181
Abstract: In-cell NMR offers great insight into the characterization of the effect of toxins and antimicrobial peptides on intact cells. However, the complexity of intact live cells remains a significant challenge for the analysis of the effect these agents have on different cellular components. Here we show that 31P solid-state NMR can be used to quantitatively characterize the dynamic behaviour of DNA within intact live bacteria. Lipids were also identified and monitored, although 31P dynamic filtering methods indicated a range of dynamic states for phospholipid headgroups. We demonstrate the usefulness of this methodology for monitoring the activity of the antibiotic icillin and the antimicrobial peptide (AMP) maculatin 1.1 (Mac1.1) against Gram-negative bacteria. Perturbations in the dynamic behaviour of DNA were observed in treated cells, which indicated additional mechanisms of action for the AMP Mac1.1 not previously reported. This work highlights the value of 31P in-cell solid-state NMR as a tool for assessing the antimicrobial activity of antibiotics and AMPs in bacterial cells.
Supramolecular Assembly of Polyphenols and Nucleic Acids by Thermal Cycling for Immune Cell Activation
Publisher: American Chemical Society (ACS)
Date: 24-08-2023
Nuclear Magnetic Resonance Study of the Peptide FRANCESSEPAROVIC
Publisher: CSIRO Publishing
Date: 2020
DOI: 10.1071/CH19357
Abstract: As an eminent ambassador of STEM and renowned NMR spectroscopist, Frances Separovic is an internationally famous name, but could it also be a valuable membrane-active peptide sequence? Her name has been used as an amino acid sequence (FS), successfully synthesised, oxidised, and put into contact with membrane models to investigate any serendipitous activity. The 3D structure of the cyclic FS was determined in dodecylphosphocholine (DPC) micelles by solution NMR spectroscopy. FS displayed a twisted bend separating a helical stretch and an unstructured segment. Using solid-state NMR spectroscopy, the effect of FS on 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and 1,2-dimyristoyl-sn-glycero-3-phosphoserine (DMPS) lipid bilayers was studied. FS did not strongly disturb the neutral membrane surface but likely inserted into their hydrophobic core without a strong effect on the lipid dynamics, while perturbation of the negatively charged membranes remained at the headgroup interface with a strong effect on the lipid dynamics. This study demonstrated that FS is a candidate for discovering potential future therapeutic activities.
Implications of peptide assemblies in amyloid diseases
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7CS00372B
Abstract: We highlight the role of molecular self-assembly in eliciting the mesoscopic and pathological properties of amyloid proteins. This knowledge is pivotal for the development of theranostics against amyloid diseases.
The antimicrobial peptide aurein 1.2 disrupts model membranes via the carpet mechanism
Publisher: Royal Society of Chemistry (RSC)
Date: 2012
DOI: 10.1039/C2CP43099A
Abstract: The membrane interactions of the antimicrobial peptide aurein 1.2 were studied using a range of biophysical techniques to determine the location and the mechanism of action in DMPC (dimyristoylphosphatidylcholine) and DMPC/DMPG (dimyristoylphosphatidylglycerol) model membranes that mimic characteristics of eukaryotic and prokaryotic membranes, respectively. Neutron reflectometry and solid-state NMR revealed subtle changes in membrane structure caused by the peptide. Quartz crystal microbalance with dissipation, vesicle dye leakage and atomic force microscopy measurements were used to investigate the global mode of peptide interaction. Aurein 1.2 displayed an enhanced interaction with the anionic DMPC/DMPG membrane while exhibiting primarily a surface interaction with both types of model membranes, which led to bilayer disruption and membrane lysis. The antimicrobial peptide interaction is consistent with the carpet mechanism for aurein 1.2 with discrete structural changes depending on the type of phospholipid membrane.
Exploring Artificial Nucleic Acid Mimicking Peptide Nanofibers
Publisher: American Chemical Society (ACS)
Date: 17-05-2023
Systematic comparison of activity and mechanism of antimicrobial peptides against nosocomial pathogens
Publisher: Elsevier BV
Date: 03-2022
DOI: 10.1016/J.EJMECH.2022.114135
Abstract: The World Health Organisation has deemed several multi-drug resistant (MDR) nosocomial bacterial pathogens to be of significant threat to human health. A stark increase in morbidity, mortality and the burden to healthcare systems around the world can be attributed to the development of resistance in these bacteria. Accordingly, alternative antimicrobial agents have been sought as an attractive means to combat MDR pathogens, with one such ex le being antimicrobial peptides (AMPs). Given the reported activity of AMPs, including Pardaxin, MSI-78, dermaseptin-PC (DMPC) and Cecropin B, it is important to understand their activities and modes of action against bacteria for further AMP design. In this study, we compared these AMPs against a panel of nosocomial bacterial pathogens, followed by detailed mechanistic studies. It was found that Pardaxin (1-22) and MSI-78 (4-20) displayed the most pronounced antimicrobial activity against the tested bacteria. The mechanistic studies by membrane permeability and molecular dynamics simulation further confirmed the strong membrane interaction and structure of Pardaxin (1-22) and MSI-78 (4-20), which contributed to their potent activity. This study demonstrated a structure and activity guidance for further design of Pardaxin (1-22) and MSI-78 (4-20) as therapeutics against MDR pathogens. The different effects of DMPC (1-19) and Cecropin B (1-21) on membrane integrity and phospholipid membrane interactions provided critical information for the rational design of next-generation analogues with specificity against either Gram-negative or Gram-positive bacteria.
Cholesterol-Dependent Cytolysins: Membrane and Protein Structural Requirements for Pore Formation
Publisher: American Chemical Society (ACS)
Date: 17-06-2019
DOI: 10.1021/ACS.CHEMREV.9B00090
Abstract: The cholesterol-dependent cytolysins (CDCs) are a family of bacterial protein toxins specifically targeting eukaryotic cells through the absolute requirement for high concentrations of cholesterol in the target cells' lipid membrane. The soluble monomeric protein secreted by the bacteria oligomerizes on the surface of the target cell, and the complex formed then undergoes a concerted structural transition that results in the creation of a multimeric protein pore. Recognition of the cholesterol-rich membrane by CDCs is a surprisingly subtle process that takes place at the interface between the membrane and surrounding aqueous environment. The structure and composition of the lipid membrane modulates the efficiency with which the protein can identify cholesterol and alters the concentration of sterol required for membrane binding. Some of the details of the interplay between protein and membrane remain to be resolved, and in this review we present a current perspective on CDC pore formation, with particular focus on the role of the lipid bilayer and cholesterol accessibility.
How do Self-Assembling Antimicrobial Lipopeptides Kill Bacteria?
Publisher: American Chemical Society (ACS)
Date: 12-2020
Investigating the Interaction of Octapeptin A3 with Model Bacterial Membranes
Publisher: American Chemical Society (ACS)
Date: 11-07-2017
Lipid composition regulates the conformation and insertion of the antimicrobial peptide maculatin 1.1
Publisher: Elsevier BV
Date: 02-2012
DOI: 10.1016/J.BBAMEM.2011.07.015
Abstract: Antimicrobial peptides interact with cell membranes and their selectivity is contingent on the nature of the constituent lipids. Eukaryotic and bacterial membranes are comprised of different proportions of a range of lipid species with different physical properties. Hence, characterisation of antimicrobial peptides with respect to the magnitude of their interactions with model membranes of different lipid types is needed. Maculatin 1.1 is a short antimicrobial peptide secreted from the skin of several Australian tree-frog species. Circular dichroism spectroscopy (CD) was used to explore the interaction of maculatin 1.1 with a wide range of model membrane systems of different head group and acyl chain characteristics. For neutral phosphatidylcholine (PC), unlike anionic phospholipids, the magnitude of the peptide interactions was dependent on the length and degree of saturation of the constituent acyl chains. Oriented circular dichroism (OCD) data indicated that helical structure was likely promoted by peptide insertion into the hydrophobic core of PC bilayers. The addition of cholesterol (30% mol/mol) tended to decrease the membrane interaction of maculatin 1.1. Anionic lipids locked maculatin 1.1 via electrostatic interactions onto the surface of oriented bilayers as seen in OCD spectra. Furthermore, increasing the membrane curvature by reducing the vesicle radii only slightly reduced the proportion of helical structure in all systems by approximately 10%. The peptide-lipid interaction was strongly dependent on both the lipid chain length and head group, which highlights the importance of the lipid composition used to mimic different cell types. This article is part of a Special Issue entitled: Membrane protein structure and function.
High-Performance Coral Reef-like Carbon Nitrides: Synthesis and Application in Photocatalysis and Heavy Metal Ion Adsorption
Publisher: American Chemical Society (ACS)
Date: 30-01-2017
Abstract: Synthesis of carbon nitrides (CN
Synthesis and secondary structure in membranes of the Bcl‐2 anti‐apoptotic domain BH4
Publisher: Wiley
Date: 09-06-2005
DOI: 10.1002/PSC.686
Abstract: Solid phase synthesis of BH4, the 26 amino-acid domain (6RTGYDNREIVMKYIHYKLSQRGYEWD31) of the anti-apoptotic Bcl-2 protein has been accomplished using Fmoc chemistry. The use of peculiar cleavage conditions provided high yields after purification such that tens to hundreds of mg could be obtained. A 15N-labelled version of the peptide could also be synthesized for NMR studies in membranes. The peptide purity was not lower than 98% as controlled by UV and MALDI-TOF mass spectrometry. The secondary structure was determined in water, trifluoroethanol (TFE) and in lipid membrane using UV circular dichroism. The peptide shows dominant beta-sheeted structures in water that convert progressively into alpha-helical features upon addition of TFE or membrane. The hipathic character of the helix suggests that the peptide might have a structure akin to those of antimicrobial peptides upon interaction with membranes.
Membrane accessibility of glutathione
Publisher: Elsevier BV
Date: 10-2015
DOI: 10.1016/J.BBAMEM.2015.07.016
Abstract: Regulation of the ion pumping activity of the Na+,K+-ATPase is crucial to the survival of animal cells. Recent evidence has suggested that the activity of the enzyme could be controlled by glutathionylation of cysteine residue 45 of the β-subunit. Crystal structures so far available indicate that this cysteine is in a transmembrane domain of the protein. Here we have analysed via fluorescence and NMR spectroscopy as well as molecular dynamics simulations whether glutathione is able to penetrate into the interior of a lipid membrane. No evidence for any penetration of glutathione into the membrane was found. Therefore, the most likely mechanism whereby the cysteine residue could become glutathionylated is via a loosening of the α-β subunit association, creating a hydrophilic passageway between them to allow access of glutathione to the cysteine residue. By such a mechanism, glutathionylation of the protein would be expected to anchor the modified cysteine residue in a hydrophilic environment, inhibiting further motion of the β-subunit during the enzyme's catalytic cycle and suppressing enzymatic activity, as has been experimentally observed. The results obtained, therefore, suggest a possible structural mechanism of how the Na+,K+-ATPase could be regulated by glutathione.
Nitroxide spin–labeled peptides for DNP‐NMR in‐cell studies
Publisher: Wiley
Date: 06-07-2019
Membrane interactions and biological activity of antimicrobial peptides from Australian scorpion
Publisher: Elsevier BV
Date: 09-2014
DOI: 10.1016/J.BBAMEM.2013.10.022
Abstract: UyCT peptides are antimicrobial peptides isolated from the venom of the Australian scorpion. The activity of the UyCT peptides against Gram positive and Gram negative bacteria and red blood cells was determined. The membrane interactions of these peptides were evaluated by dye release (DR) of the fluorophore calcein from liposomes and isothermal titration calorimetry (ITC) and their secondary structure was determined by circular dichroism (CD). Three different lipid systems were used to mimic red blood cells, Escherichia coli and Staphylococcus aureus membranes. UyCT peptides exhibited broad spectrum antimicrobial activity with low MIC for S. aureus and multi-drug resistant Gram negative strains. Peptide combinations showed some synergy enhancing their potency but not hemolytic activity. The UyCT peptides adopted a helical structure in lipid environments and DR results confirmed that the mechanism of action is by disrupting the membrane. ITC data indicated that UyCT peptides preferred prokaryotic rather than eukaryotic membranes. The overall results suggest that UyCT peptides could be pharmaceutical leads for the treatment of Gram negative multiresistant bacterial infections, especially against Acinetobacter baumanni, and candidates for peptidomimetics to enhance their potency and minimize hemolysis. This article is part of a Special Issue entitled: Interfacially Active Peptides and Proteins. Guest Editors: William C. Wimley and Kalina Hristova.
Transient RNA Interactions Leave a Covalent Imprint on a Viral Capsid Protein
Publisher: American Chemical Society (ACS)
Date: 05-05-2022
DOI: 10.1021/JACS.1C12439
Antimicrobial Peptide Structures: From Model Membranes to Live Cells
Publisher: Wiley
Date: 30-11-2018
Abstract: The rise in antibiotic resistance has led to a renewed interest in antimicrobial peptides (AMPs) that target membranes. The mode of action of AMPs involves the disruption of the lipid bilayer and leads to growth inhibition and death of the bacteria. However, details at the molecular level of how these peptides kill bacteria and the reasons for the observed differences in selectivity remain unclear. Structural information is crucial for defining the molecular mechanism by which these peptides recognize, self-assemble and interact with a particular lipid membrane. Solid-state NMR is a non-invasive technique that allows the study of the structural details of lipid-peptide and peptide-peptide interactions. Following on from studies of antibiotic and lytic peptides, gramicidin A and melittin, respectively, we investigated maculatin 1.1, an AMP from the skin of Australian tree frogs that acts against Gram-positive bacteria. By using perdeuterated phospholipids and specifically labelled peptides,
Pro-apoptotic bax-α1 synthesis and evidence for β-sheet to α-helix conformational change as triggered by negatively charged lipid membranes
Publisher: Wiley
Date: 2007
DOI: 10.1002/PSC.803
Abstract: Solid phase synthesis of Bax-alpha1, the 25 amino acids domain (14TSSEQIMKTGALLLQGFIQDRAGRM38) of the pro-apoptotic Bax protein has been accomplished using Fmoc chemistry. A new fast and harmless protocol is described for complete TFA removal from the purified peptide powder leading to a final purity greater than 98% as controlled by 19F-NMR, UV and MALDI-TOF mass spectrometry. Secondary structure was determined in various solution and membrane media using UV Circular Dichroism. In water solution, Bax-alpha1 is present as a mixture of beta-sheet and unstructured (random coil) conformations. A marked change from beta-sheet to alpha-helix secondary structures is observed upon interaction with negatively charged phospholipids vesicles whereas neutral lipid membranes have no significant effect on the aqueous peptide conformation. Results are discussed in terms of Bax binding to mitochondrial membranes.
Frontispiece: Antimicrobial Peptide Structures: From Model Membranes to Live Cells
Publisher: Wiley
Date: 09-01-2018
The membrane activity of the antimicrobial peptide caerin 1.1 is pH dependent
Publisher: Elsevier BV
Date: 03-2023
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