Structural insights into the interaction of antifungal peptides and ergosterol containing fungal membrane

Mohid, Sk Abdul; Biswas, Karishma; Won, TaeJun; Mallela, Lakshmi Sruthi; Gucchait, Arin; Butzke, Lena; Sarkar, Riddhiman; Barkham, Timothy; Reif, Bernd; Leipold, Enrico; Roy, Sanhita; Misra, Anup Kumar; Lakshminarayanan, Rajamani; Lee, Dong‐Kuk; Bhunia, Anirban

doi:10.1016/j.bbamem.2022.183996

Cited by 11 publications

(5 citation statements)

References 52 publications

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“…This yield is more than three times higher than the yield of GUVs obtained when ULGT agarose and PVA are used as the assisting compounds. This result is notable since ULGT agarose and PVA are both used extensively in the literature, − whereas, as far as we know, we are the first to report the use of LGT agarose. Our results are general.…”

Section: Resultsmentioning

confidence: 81%

Osmotic Pressure Enables High-Yield Assembly of Giant Vesicles in Solutions of Physiological Ionic Strengths

2023

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Giant unilamellar vesicles (GUVs) are micrometer-scale minimal cellular mimics that are useful for bottom-up synthetic biology and drug delivery. Unlike assembly in low-salt solutions, assembly of GUVs in solutions with ionic concentrations of 100–150 mM Na/KCl (salty solutions) is challenging. Chemical compounds deposited on the substrate or incorporated into the lipid mixture could assist in the assembly of GUVs. Here, we investigate quantitatively the effects of temperature and chemical identity of six polymeric compounds and one small molecule compound on the molar yields of GUVs composed of three different lipid mixtures using high-resolution confocal microscopy and large data set image analysis. All the polymers moderately increased the yields of GUVs either at 22 or 37 °C, whereas the small molecule compound was ineffective. Low-gelling temperature agarose is the singular compound that consistently produces yields of GUVs of greater than 10%. We propose a free energy model of budding to explain the effects of polymers in assisting the assembly of GUVs. The osmotic pressure exerted on the membranes by the dissolved polymer balances the increased adhesion between the membranes, thus reducing the free energy for bud formation. Data obtained by modulating the ionic strength and ion valency of the solution shows that the evolution of the yield of GUVs supports our model’s prediction. In addition, polymer-specific interactions with the substrate and the lipid mixture affects yields. The uncovered mechanistic insights provide a quantitative experimental and theoretical framework to guide future studies. Additionally, this work shows a facile means for obtaining GUVs in solutions of physiological ionic strengths.

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Section: Resultsmentioning

confidence: 81%

Osmotic Pressure Enables High-Yield Assembly of Giant Vesicles in Solutions of Physiological Ionic Strengths

2023

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“…Analysing this interaction might shed light onto the contribution of proteins to the overall fluidity and stability of lipid bilayers. Therefore, in order to study the interaction of Hsp12 with lipid, in vitro LUVs were made out of DMPG lipid as described in Mohid et al ( 2022 ) and detailed in Method S4 .…”

Section: Methodsmentioning

confidence: 99%

“…Therefore, in order to study the interaction of Hsp12 with lipid, in vitro LUVs were made out of DMPG lipid as described in Mohid et al (2022) and detailed in Method S4.…”

Section: Preparation Of Dmpg Luvsmentioning

confidence: 99%

Insight into the biochemical and cell biological function of an intrinsically unstructured heat shock protein, Hsp12 of Ustilago maydis

Mitra

Bhakta

Kar

et al. 2023

Molecular Plant Pathology

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Small heat shock proteins (sHsps) play diverse roles in the stress response and maintenance of cellular functions. The Ustilago maydis genome codes for few sHsps. Among these, Hsp12 has previously been demonstrated to be involved in the pathogenesis of the fungus by our group. In the present study we further investigated the biological function of the protein in the pathogenic development of U. maydis. Analysis of the primary amino acid sequence of Hsp12 in combination with spectroscopic methods to analyse secondary protein structures revealed an intrinsically disordered nature of the protein. We also carried out detailed analysis on the protein aggregation prevention activity associated with Hsp12. Our data suggest Hsp12 has trehalose‐dependent protein aggregation prevention activity. Through assaying the interaction of Hsp12 with lipid membranes in vitro we also showed the ability of U. maydis Hsp12 to induce stability in lipid vesicles. U. maydis hsp12 deletion mutants exhibited defects in the endocytosis process and delayed completion of the pathogenic life cycle. Therefore, U. maydis Hsp12 contributes to the pathogenic development of the fungus through its ability to relieve proteotoxic stress during infection as well as its membrane‐stabilizing function.

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“…Ergosterol biosynthesis is specific to the fungi and some algae (Darnet et al., 2021 ). This sterol is not only needed for cell membrane fluidity but may also play an important role in glycerol efflux and survival of S. cerevisiae cells during osmotic downshock and acts as a fungal hormone that stimulates growth and proliferation (Figure 1D ; Jorda & Puig, 2020 ; Mohid et al, 2022 ; Szomek et al, 2022 ; Toh et al, 2001 ). Sterol profiling has been one of the methods used to identify the targets of azole‐based compounds and other types of ergosterol biosynthesis inhibitors (Müller et al, 2017 ).…”

Section: Analysis Of Fungal Metabolitesmentioning

confidence: 99%

Modes‐of‐action of antifungal compounds: Stressors and (target‐site‐specific) toxins, toxicants, or toxin–stressors

Noël

Hallsworth

Gelhaye

et al. 2023

Microbial Biotechnology

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Fungi and antifungal compounds are relevant to the United Nation's Sustainable Development Goals. However, the modes‐of‐action of antifungals—whether they are naturally occurring substances or anthropogenic fungicides—are often unknown or are misallocated in terms of their mechanistic category. Here, we consider the most effective approaches to identifying whether antifungal substances are cellular stressors, toxins/toxicants (that are target‐site‐specific), or have a hybrid mode‐of‐action as toxin–stressors (that induce cellular stress yet are target‐site‐specific). This newly described ‘toxin–stressor’ category includes some photosensitisers that target the cell membrane and, once activated by light or ultraviolet radiation, cause oxidative damage. We provide a glossary of terms and a diagrammatic representation of diverse types of stressors, toxic substances, and toxin–stressors, a classification that is pertinent to inhibitory substances not only for fungi but for all types of cellular life. A decision‐tree approach can also be used to help differentiate toxic substances from cellular stressors (Curr Opin Biotechnol 2015 33: 228–259). For compounds that target specific sites in the cell, we evaluate the relative merits of using metabolite analyses, chemical genetics, chemoproteomics, transcriptomics, and the target‐based drug‐discovery approach (based on that used in pharmaceutical research), focusing on both ascomycete models and the less‐studied basidiomycete fungi. Chemical genetic methods to elucidate modes‐of‐action currently have limited application for fungi where molecular tools are not yet available; we discuss ways to circumvent this bottleneck. We also discuss ecologically commonplace scenarios in which multiple substances act to limit the functionality of the fungal cell and a number of as‐yet‐unresolved questions about the modes‐of‐action of antifungal compounds pertaining to the Sustainable Development Goals.

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Structural insights into the interaction of antifungal peptides and ergosterol containing fungal membrane

Cited by 11 publications

References 52 publications

Osmotic Pressure Enables High-Yield Assembly of Giant Vesicles in Solutions of Physiological Ionic Strengths

Osmotic Pressure Enables High-Yield Assembly of Giant Vesicles in Solutions of Physiological Ionic Strengths

Insight into the biochemical and cell biological function of an intrinsically unstructured heat shock protein, Hsp12 of Ustilago maydis

Modes‐of‐action of antifungal compounds: Stressors and (target‐site‐specific) toxins, toxicants, or toxin–stressors

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