Pub Date : 2026-01-01Epub Date: 2025-10-10DOI: 10.1016/j.bbamem.2025.184475
Thomas Heimburg , Holger Ebel , Peter Grabitz , Julia Preu , Yue Wang
During the melting transition of dimyristoyl phosphatidylglycerol (DMPG), the order of the lipid chains and the three-dimensional, vesicular structural arrangement change simultaneously. These changes result in peculiar heat capacity profiles extended over a broad temperature range with several -maxima. Here, we present calorimetric, viscosity, and volume expansion coefficient data at various ionic strengths and charges. We propose a simple theory that explains the calorimetric data in terms of the coexistence of two membrane geometries, both of which can melt. During the transition, the equilibrium between these two geometries changes cooperatively. This equilibrium depends on the interactions between the membranes and the solvent, on the membrane’s charge and the ionic strength of the buffer. Solvent interactions also contribute to the volume change of the membrane phases. Unlike uncharged membranes, we find that enthalpy changes are no longer proportional to volume changes. Therefore, the pressure dependence of the calorimetric profiles differs from that of uncharged membranes. Our theory explains the pressure dependence of calorimetric profiles qualitatively and quantitatively. Furthermore, we demonstrate that the same theory can be used to describe pretransition and ripple formation in phosphatidylcholines. A key takeaway from this article is that solvent molecules (e.g., HO) are part of the membrane and, in the case of DMPG, water cannot be considered a separate phase.
{"title":"The relationship between the structural transitions of DMPG membranes and the melting process, and their interaction with water","authors":"Thomas Heimburg , Holger Ebel , Peter Grabitz , Julia Preu , Yue Wang","doi":"10.1016/j.bbamem.2025.184475","DOIUrl":"10.1016/j.bbamem.2025.184475","url":null,"abstract":"<div><div>During the melting transition of dimyristoyl phosphatidylglycerol (DMPG), the order of the lipid chains and the three-dimensional, vesicular structural arrangement change simultaneously. These changes result in peculiar heat capacity profiles extended over a broad temperature range with several <span><math><msub><mrow><mi>c</mi></mrow><mrow><mi>p</mi></mrow></msub></math></span>-maxima. Here, we present calorimetric, viscosity, and volume expansion coefficient data at various ionic strengths and charges. We propose a simple theory that explains the calorimetric data in terms of the coexistence of two membrane geometries, both of which can melt. During the transition, the equilibrium between these two geometries changes cooperatively. This equilibrium depends on the interactions between the membranes and the solvent, on the membrane’s charge and the ionic strength of the buffer. Solvent interactions also contribute to the volume change of the membrane phases. Unlike uncharged membranes, we find that enthalpy changes are no longer proportional to volume changes. Therefore, the pressure dependence of the calorimetric profiles differs from that of uncharged membranes. Our theory explains the pressure dependence of calorimetric profiles qualitatively and quantitatively. Furthermore, we demonstrate that the same theory can be used to describe pretransition and ripple formation in phosphatidylcholines. A key takeaway from this article is that solvent molecules (e.g., H<span><math><msub><mrow></mrow><mrow><mn>2</mn></mrow></msub></math></span>O) are part of the membrane and, in the case of DMPG, water cannot be considered a separate phase.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1868 1","pages":"Article 184475"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145278921","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-01-01Epub Date: 2025-10-05DOI: 10.1016/j.bbamem.2025.184470
Joseph C. Iovine , Benjamin T. Garrett , Nathan N. Alder
Styrene-maleic acid (SMA) copolymers are powerful tools for the detergent-free solubilization of biological membranes. Yet the influence of specific lipids on SMA activity remains an open question. Here, we examined the effects of the mitochondria-specific phospholipid cardiolipin on SMA-mediated membrane solubilization and its ability to form SMA-bound nanodiscs. To this end, we prepared a series of model membranes with cardiolipin and other test lipids with comparable surface charge and lateral packing characteristics. Using multiple independent experimental approaches, we found that cardiolipin inhibited SMA solubilization. Our results indicate that this effect was not attributable to headgroup charge effects, but related to cardiolipin-induced increase in lateral packing pressure at the interfacial region. Reduction of this lateral packing pressure using bilayer-active alcohols partially restored SMA solubilization. Our results highlight the importance of lipid geometry and packing in SMA nanodisc formation and could help guide the design of copolymers tailored to specific membranes.
{"title":"Lateral lipid packing governs bilayer solubilization by styrene-maleic acid copolymers: a case study with cardiolipin-containing membranes","authors":"Joseph C. Iovine , Benjamin T. Garrett , Nathan N. Alder","doi":"10.1016/j.bbamem.2025.184470","DOIUrl":"10.1016/j.bbamem.2025.184470","url":null,"abstract":"<div><div>Styrene-maleic acid (SMA) copolymers are powerful tools for the detergent-free solubilization of biological membranes. Yet the influence of specific lipids on SMA activity remains an open question. Here, we examined the effects of the mitochondria-specific phospholipid cardiolipin on SMA-mediated membrane solubilization and its ability to form SMA-bound nanodiscs. To this end, we prepared a series of model membranes with cardiolipin and other test lipids with comparable surface charge and lateral packing characteristics. Using multiple independent experimental approaches, we found that cardiolipin inhibited SMA solubilization. Our results indicate that this effect was not attributable to headgroup charge effects, but related to cardiolipin-induced increase in lateral packing pressure at the interfacial region. Reduction of this lateral packing pressure using bilayer-active alcohols partially restored SMA solubilization. Our results highlight the importance of lipid geometry and packing in SMA nanodisc formation and could help guide the design of copolymers tailored to specific membranes.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1868 1","pages":"Article 184470"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145243668","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-01-01Epub Date: 2025-10-25DOI: 10.1016/j.bbamem.2025.184478
Paulo Henrique Lima do Nascimento , Kevin Figueiredo dos Santos , Cristiano Giordani , Joelle Mergola-Greef , Marcel Jaspars , Luciano Caseli
Patellamides are cyclic pseudo-octapeptides derived from marine cyanobacteria with promising selective cytotoxic, antimicrobial, and neuroprotective activities. While their biological potential is increasingly recognized, the mechanisms underlying their interaction with lipid membranes remain poorly understood. In this study, we investigated the interfacial behavior of patellamide D using Langmuir monolayers composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphoserine (DPPS), which model the outer leaflets of healthy and cancer cell membranes, respectively. Surface pressure–area isotherms, compressional modulus analysis, surface potential measurements, Brewster angle microscopy, Polarization-modulated infrared reflection-absorption spectroscopy, and interfacial rheology were employed to elucidate peptide–lipid interactions. Patellamide exhibited a lipid-specific condensing effect and induced subtle reorganization within the monolayers, particularly in anionic DPPS films. Despite these interactions, the compressional and viscoelastic properties of the monolayers were largely preserved, suggesting stable incorporation of the peptide without compromising film integrity. These findings reveal that patellamide can modulate lipid packing and interface properties in a selective and controlled manner. Such behavior underscores its potential in the design of membrane-active therapeutic agents and lipid-based drug delivery systems.
{"title":"Patellamide–lipid interactions at the air–water interface: Biophysical insights into membrane modulation","authors":"Paulo Henrique Lima do Nascimento , Kevin Figueiredo dos Santos , Cristiano Giordani , Joelle Mergola-Greef , Marcel Jaspars , Luciano Caseli","doi":"10.1016/j.bbamem.2025.184478","DOIUrl":"10.1016/j.bbamem.2025.184478","url":null,"abstract":"<div><div>Patellamides are cyclic pseudo-octapeptides derived from marine cyanobacteria with promising selective cytotoxic, antimicrobial, and neuroprotective activities. While their biological potential is increasingly recognized, the mechanisms underlying their interaction with lipid membranes remain poorly understood. In this study, we investigated the interfacial behavior of patellamide D using Langmuir monolayers composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphoserine (DPPS), which model the outer leaflets of healthy and cancer cell membranes, respectively. Surface pressure–area isotherms, compressional modulus analysis, surface potential measurements, Brewster angle microscopy, Polarization-modulated infrared reflection-absorption spectroscopy, and interfacial rheology were employed to elucidate peptide–lipid interactions. Patellamide exhibited a lipid-specific condensing effect and induced subtle reorganization within the monolayers, particularly in anionic DPPS films. Despite these interactions, the compressional and viscoelastic properties of the monolayers were largely preserved, suggesting stable incorporation of the peptide without compromising film integrity. These findings reveal that patellamide can modulate lipid packing and interface properties in a selective and controlled manner. Such behavior underscores its potential in the design of membrane-active therapeutic agents and lipid-based drug delivery systems.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1868 1","pages":"Article 184478"},"PeriodicalIF":2.5,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145413688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-09-22DOI: 10.1016/j.bbamem.2025.184459
Brandon S. Veron , Kyle Lethcoe , Robert O. Ryan
The SpyCatcher/SpyTag system represents a unique technology that allows facile conjugation of proteins via formation of a covalent isopeptide bond between the 113 residue SpyCatcher protein and a 16 residue SpyTag peptide. Herein this technology was adapted to incorporate miniature bilayer membranes, termed nanodisks (ND). Fusion proteins comprised of apolipoprotein (apo) A-I/SpyTag peptide and SpyCatcher/maltose binding protein (MBP), respectively, were expressed and purified. Upon incubation of apoA-I:SpyTag fusion protein with SpyCatcher:MBP fusion protein, a covalent adduct was formed. ApoA-I:SpyTag formulated into ND particles with cardiolipin (CL) or phosphatidylcholine retained the ability to form an adduct with SpyCatcher:MBP. This adduct was then immobilized on amylose agarose resin beads through a binding interaction with the MBP component. Upon incubation of cytochrome c with immobilized CL ND, but not with phosphatidylcholine ND, cytochrome c binding occurred. When immobilized cytochrome c CL ND were incubated with buffer containing CaCl2, cytochrome c dissociated and was recovered in the supernatant fraction obtained after pelleting the amylose agarose beads. Subsequent incubation of the amylose agarose beads with 10 mM maltose revealed that nearly all of the cytochrome c had been released from the beads. The data are consistent with the known ability of calcium to form an ionic interaction with the two negatively charged phosphates in the polar head group of CL. Given the number of ligand-membrane interactions that occur in nature, immobilized ND provide a novel means to probe them.
{"title":"Immobilized nanodisks for study of ligand binding interactions","authors":"Brandon S. Veron , Kyle Lethcoe , Robert O. Ryan","doi":"10.1016/j.bbamem.2025.184459","DOIUrl":"10.1016/j.bbamem.2025.184459","url":null,"abstract":"<div><div>The SpyCatcher/SpyTag system represents a unique technology that allows facile conjugation of proteins via formation of a covalent isopeptide bond between the 113 residue SpyCatcher protein and a 16 residue SpyTag peptide. Herein this technology was adapted to incorporate miniature bilayer membranes, termed nanodisks (ND). Fusion proteins comprised of apolipoprotein (apo) A-I/SpyTag peptide and SpyCatcher/maltose binding protein (MBP), respectively, were expressed and purified. Upon incubation of apoA-I:SpyTag fusion protein with SpyCatcher:MBP fusion protein, a covalent adduct was formed. ApoA-I:SpyTag formulated into ND particles with cardiolipin (CL) or phosphatidylcholine retained the ability to form an adduct with SpyCatcher:MBP. This adduct was then immobilized on amylose agarose resin beads through a binding interaction with the MBP component. Upon incubation of cytochrome <em>c</em> with immobilized CL ND, but not with phosphatidylcholine ND, cytochrome <em>c</em> binding occurred. When immobilized cytochrome <em>c</em> CL ND were incubated with buffer containing CaCl<sub>2</sub>, cytochrome <em>c</em> dissociated and was recovered in the supernatant fraction obtained after pelleting the amylose agarose beads. Subsequent incubation of the amylose agarose beads with 10 mM maltose revealed that nearly all of the cytochrome <em>c</em> had been released from the beads. The data are consistent with the known ability of calcium to form an ionic interaction with the two negatively charged phosphates in the polar head group of CL. Given the number of ligand-membrane interactions that occur in nature, immobilized ND provide a novel means to probe them.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184459"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145118696","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-08-27DOI: 10.1016/j.bbamem.2025.184446
B.Z. Favarin , N. Nassif , T. Azaïs , J. Guignier , S. Mebarek , R. Buchet , J.L. Millán , A.P. Ramos , A.J. Costa-Filho , P. Ciancaglini
Skeletal and dental mineralization relies on a precisely regulated sequence of events culminating in apatite deposition onto collagen fibrils. Matrix vesicles (MVs), extracellular vesicles released by mineralization-competent cells, play a pivotal role in this process through the catalytic activity of alkaline phosphatase (TNAP). The lipid composition of MVs, particularly phosphatidylserine (PS)-calcium complexes, facilitates the nucleation of amorphous calcium phosphate and apatite formation. However, the interplay between the TNAP structure, the lipid membrane environment, and its enzymatic activity remains incompletely understood.
Biomimetic models of MVs, as proteoliposomes made with dipalmitoylphosphatidylcholine (DPPC) and various TNAP mutants, were used to investigate the TNAP's activity and mineralization potential. Molecular docking and site-directed mutagenesis revealed that specific cysteine substitutions near TNAP's catalytic and anchoring sites influence structural stability, enzymatic activity, and incorporation into lipid bilayers. Notably, TNAP mutants S221C and P307C exhibited enhanced catalytic efficiency in DPPC liposomes, while A420C showed reduced activity due to steric hindrance near the catalytic site. Solid-state NMR and cryo-TEM analyses confirmed hydroxyapatite formation, with significant contributions from lipid-anchored TNAP to the mineralization process.
These findings highlight the critical influence of the lipid environment on TNAP's functional properties and provide insights into the mechanisms governing biomineralization and related pathologies, including hypophosphatasia associated with various TNAP mutations. The study underscores the importance of ATP and pyrophosphate hydrolysis by TNAP in modulating apatite formation and reveals the role of specific TNAP mutations in regulating enzymatic activity, stability, and mineral propagation. Understanding these interactions could lead to alternate therapeutic strategies in treatment and regenerative medicine.
{"title":"Modulation of TNAP activity and apatite formation in biomimetic matrix vesicles studied by 31P solid-state NMR","authors":"B.Z. Favarin , N. Nassif , T. Azaïs , J. Guignier , S. Mebarek , R. Buchet , J.L. Millán , A.P. Ramos , A.J. Costa-Filho , P. Ciancaglini","doi":"10.1016/j.bbamem.2025.184446","DOIUrl":"10.1016/j.bbamem.2025.184446","url":null,"abstract":"<div><div>Skeletal and dental mineralization relies on a precisely regulated sequence of events culminating in apatite deposition onto collagen fibrils. Matrix vesicles (MVs), extracellular vesicles released by mineralization-competent cells, play a pivotal role in this process through the catalytic activity of alkaline phosphatase (TNAP). The lipid composition of MVs, particularly phosphatidylserine (PS)-calcium complexes, facilitates the nucleation of amorphous calcium phosphate and apatite formation. However, the interplay between the TNAP structure, the lipid membrane environment, and its enzymatic activity remains incompletely understood.</div><div>Biomimetic models of MVs, as proteoliposomes made with dipalmitoylphosphatidylcholine (DPPC) and various TNAP mutants, were used to investigate the TNAP's activity and mineralization potential. Molecular docking and site-directed mutagenesis revealed that specific cysteine substitutions near TNAP's catalytic and anchoring sites influence structural stability, enzymatic activity, and incorporation into lipid bilayers. Notably, TNAP mutants S221C and P307C exhibited enhanced catalytic efficiency in DPPC liposomes, while A420C showed reduced activity due to steric hindrance near the catalytic site. Solid-state NMR and cryo-TEM analyses confirmed hydroxyapatite formation, with significant contributions from lipid-anchored TNAP to the mineralization process.</div><div>These findings highlight the critical influence of the lipid environment on TNAP's functional properties and provide insights into the mechanisms governing biomineralization and related pathologies, including hypophosphatasia associated with various TNAP mutations. The study underscores the importance of ATP and pyrophosphate hydrolysis by TNAP in modulating apatite formation and reveals the role of specific TNAP mutations in regulating enzymatic activity, stability, and mineral propagation. Understanding these interactions could lead to alternate therapeutic strategies in treatment and regenerative medicine.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184446"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144940771","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-09-11DOI: 10.1016/j.bbamem.2025.184458
Valentina A. Iunusova , Nikita A. Orlov , Oksana V. Nekrasova , Alexey V. Feofanov , Alexander A. Vassilevski , Alexey I. Kuzmenkov
KcsA, a potassium channel from Streptomyces lividans, is one of the most extensively studied transmembrane proteins. Despite significant research in structural biology, relatively few ligands of KcsA have been identified. One such ligand is Hui1, an artificial peptide derived from a phage display screening using a combinatorial library constructed relying on several sea anemone toxins. In this study, we engineered a fluorescent probe by fusing Hui1 with enhanced green fluorescent protein (eGFP), creating the first fluorescence-based tool to visualize prokaryotic ion channels. The eGFP–Hui1 chimera was successfully produced in Escherichia coli and purified using chromatographic techniques. Our study revealed a direct interaction between KcsA, also recombinantly expressed in E. coli, and the fluorescent chimera. Furthermore, we demonstrated that both Hui1 and tetraethylammonium can effectively displace the chimera from its complex with KcsA, confirming the specificity of the binding interaction. This approach opens new avenues for pharmacological and structural investigations, including the development of novel antimicrobial agents and high-throughput ligand screening.
{"title":"Targeting prokaryotic ion channel by a chimera of fluorescent protein and artificial peptide toxin","authors":"Valentina A. Iunusova , Nikita A. Orlov , Oksana V. Nekrasova , Alexey V. Feofanov , Alexander A. Vassilevski , Alexey I. Kuzmenkov","doi":"10.1016/j.bbamem.2025.184458","DOIUrl":"10.1016/j.bbamem.2025.184458","url":null,"abstract":"<div><div>KcsA, a potassium channel from <em>Streptomyces lividans</em>, is one of the most extensively studied transmembrane proteins. Despite significant research in structural biology, relatively few ligands of KcsA have been identified. One such ligand is Hui1, an artificial peptide derived from a phage display screening using a combinatorial library constructed relying on several sea anemone toxins. In this study, we engineered a fluorescent probe by fusing Hui1 with enhanced green fluorescent protein (eGFP), creating the first fluorescence-based tool to visualize prokaryotic ion channels. The eGFP–Hui1 chimera was successfully produced in <em>Escherichia coli</em> and purified using chromatographic techniques. Our study revealed a direct interaction between KcsA, also recombinantly expressed in <em>E. coli</em>, and the fluorescent chimera. Furthermore, we demonstrated that both Hui1 and tetraethylammonium can effectively displace the chimera from its complex with KcsA, confirming the specificity of the binding interaction. This approach opens new avenues for pharmacological and structural investigations, including the development of novel antimicrobial agents and high-throughput ligand screening.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184458"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145058265","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-08-19DOI: 10.1016/j.bbamem.2025.184445
Elena T. Aliper , Roman G. Efremov
The coronavirus spike protein, the key entity effectuating membrane fusion, cannot exist without membrane-active fragments. In addition to fusion peptides, among such domains are HR1 and HR2. Crucial to the spike's refolding and membrane fusion, they are believed to both interact with each other and bind to the membranes that are merged. To elucidate HR2's precise role in this process, an understanding of its structure and behaviour is required. Here, we used various computational approaches to study SARS-CoV-2 spike HR2's (1163-1211) interaction with membranes in the context within which it operates in live spike. During simulations with model bilayers, HR2 remained hugely unresponsive to the presence of a membrane, however, when extended to include the transmembrane domain (TMD) (1212-1234) and/or membrane-active preHR2 fragment (1147-1161), HR2’s binding to model bilayers was markedly enhanced. The trimeric coiled-coil of HR2 does not dissociate either on its own or with added TMD and/or preHR2. Molecular hydrophobicity potential (MHP) mapping showed that HR2's central part possesses a tilted oblique-oriented motif characteristic of “textbook” membrane-active peptides, albeit flanked by highly hydrophilic fragments. A truncated HR2 only encompassing this motif had a greater affinity for membranes, suggesting HR2 has a modular structure with a membrane-active segment masked by flanking regions and might be potentiated by HR2-adjacent domains and other factors coming into play after the spike gets enzymatically cleaved. Such a modular structure may have evolved for HR2's membrane activity to be regulated very subtly and “switched on” at precisely the right moment during viral fusion.
{"title":"The coronavirus spike HR2 domain: An obscure player entering the limelight during membrane fusion?","authors":"Elena T. Aliper , Roman G. Efremov","doi":"10.1016/j.bbamem.2025.184445","DOIUrl":"10.1016/j.bbamem.2025.184445","url":null,"abstract":"<div><div>The coronavirus spike protein, the key entity effectuating membrane fusion, cannot exist without membrane-active fragments. In addition to fusion peptides, among such domains are HR1 and HR2. Crucial to the spike's refolding and membrane fusion, they are believed to both interact with each other and bind to the membranes that are merged. To elucidate HR2's precise role in this process, an understanding of its structure and behaviour is required. Here, we used various computational approaches to study SARS-CoV-2 spike HR2's (1163-1211) interaction with membranes in the context within which it operates in live spike. During simulations with model bilayers, HR2 remained hugely unresponsive to the presence of a membrane, however, when extended to include the transmembrane domain (TMD) (1212-1234) and/or membrane-active preHR2 fragment (1147-1161), HR2’s binding to model bilayers was markedly enhanced. The trimeric coiled-coil of HR2 does not dissociate either on its own or with added TMD and/or preHR2. Molecular hydrophobicity potential (MHP) mapping showed that HR2's central part possesses a tilted oblique-oriented motif characteristic of “textbook” membrane-active peptides, albeit flanked by highly hydrophilic fragments. A truncated HR2 only encompassing this motif had a greater affinity for membranes, suggesting HR2 has a modular structure with a membrane-active segment masked by flanking regions and might be potentiated by HR2-adjacent domains and other factors coming into play after the spike gets enzymatically cleaved. Such a modular structure may have evolved for HR2's membrane activity to be regulated very subtly and “switched on” at precisely the right moment during viral fusion.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184445"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144933285","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-09-11DOI: 10.1016/j.bbamem.2025.184451
Mark J. Arcario , Vikram Dalal , David Fan , Fong-Fu Hsu , Wayland W.L. Cheng
Nanodiscs, soluble membrane mimetics composed of an amphipathic membrane scaffold protein encircling a lipid bilayer, are widely used in biophysical and structural studies of membrane proteins. Because many membrane proteins are responsive to their membrane environment, through specific protein–lipid interactions and bulk membrane shape and structure, it is important to understand the properties of lipid bilayers contained within nanodiscs in order to interpret studies using this technology. Nanodiscs are known to alter lipid properties, such as membrane thickness and melting temperature, and interactions with the nanodisc rim have been hypothesized to produce local perturbations in lipid structure and dynamics. Larger nanodiscs should compensate for this effect with a larger unperturbed area. To test this hypothesis, we examined the lipid bilayer properties of several lipids (DMPC, DPPC, POPC, DSPC) and soy polar lipid in circularized nanodiscs of 11 nm to 50 nm diameter using the environmentally-sensitive fluorophore, Laurdan. In nanodiscs containing a single lipid type, as nanodisc size increased, lipid packing, melting temperature, and cooperativity better approximated the properties of that lipid in large unilamellar vesicles (LUVs). In spNW50 (50 nm nanodisc), the lipid packing and melting temperature were indistinguishable from LUVs. However, nanodiscs containing soy polar lipids did not follow this trend suggesting that complex lipid mixtures may produce preferential incorporation of lipids into the nanodisc or nonhomogeneous distribution of lipids within the nanodisc.
{"title":"Examining the thermotropic properties of large circularized nanodiscs","authors":"Mark J. Arcario , Vikram Dalal , David Fan , Fong-Fu Hsu , Wayland W.L. Cheng","doi":"10.1016/j.bbamem.2025.184451","DOIUrl":"10.1016/j.bbamem.2025.184451","url":null,"abstract":"<div><div>Nanodiscs, soluble membrane mimetics composed of an amphipathic membrane scaffold protein encircling a lipid bilayer, are widely used in biophysical and structural studies of membrane proteins. Because many membrane proteins are responsive to their membrane environment, through specific protein–lipid interactions and bulk membrane shape and structure, it is important to understand the properties of lipid bilayers contained within nanodiscs in order to interpret studies using this technology. Nanodiscs are known to alter lipid properties, such as membrane thickness and melting temperature, and interactions with the nanodisc rim have been hypothesized to produce local perturbations in lipid structure and dynamics. Larger nanodiscs should compensate for this effect with a larger unperturbed area. To test this hypothesis, we examined the lipid bilayer properties of several lipids (DMPC, DPPC, POPC, DSPC) and soy polar lipid in circularized nanodiscs of 11 nm to 50 nm diameter using the environmentally-sensitive fluorophore, Laurdan. In nanodiscs containing a single lipid type, as nanodisc size increased, lipid packing, melting temperature, and cooperativity better approximated the properties of that lipid in large unilamellar vesicles (LUVs). In spNW50 (50 nm nanodisc), the lipid packing and melting temperature were indistinguishable from LUVs. However, nanodiscs containing soy polar lipids did not follow this trend suggesting that complex lipid mixtures may produce preferential incorporation of lipids into the nanodisc or nonhomogeneous distribution of lipids within the nanodisc.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184451"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145046028","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-09-08DOI: 10.1016/j.bbamem.2025.184450
Éder Jéferson Souza Cardoso , Ellyêssa Nascimento Borges , Kleber Santiago Freitas e Silva , Rodrigo Saar Gomes , Fabiana Taniguchi Boni , Jhon Kennedy Alves Pereira , Paula Maria Alexandre Mangoba , Luciano Morais Lião , Ricardo Menegatti , Antonio Alonso
Three antileishmanial compounds incorporating a butylated hydroxytoluene (BHT) moiety and an acrylate-based Michael acceptor scaffold were rationally designed from the lead structures LQFM064 and LQFM332, which feature a chalcone-derived core. Their activities against Leishmania (L.) amazonensis were evaluated. Acrylate derivatives (5), (6), and (7) displayed IC50 values comparable to miltefosine, while showing markedly lower cytotoxicity toward J774.A1 and differentiated THP-1 macrophages, along with reduced hemolytic potential. Spin-label electron paramagnetic resonance (EPR) spectroscopy revealed that treatment with these compounds induces membrane rigidity after 24 h in a concentration-dependent manner. This effect is unlikely due to direct membrane interaction, as it does not occur after short incubations or at low concentrations, suggesting a correlation with oxidative stress, such as lipid peroxidation and/or protein oxidation, likely triggered by elevated reactive oxygen species (ROS) production. In contrast, no oxidative stress-induced membrane rigidity was detected in uninfected macrophages, suggesting that nitric oxide production may mitigate oxidative damage in these cells. However, significant membrane rigidity was observed in Leishmania-infected macrophages at concentrations slightly above the IC50 for amastigotes, indicating that the compounds may selectively target infected macrophages. Additionally, compound (5) exhibited moderate cytotoxicity in the rapidly proliferating J774.A1 macrophage line but displayed very low cytotoxicity in differentiated, non-proliferative THP-1 macrophages. Overall, this study suggests that the primary mechanisms underlying the antileishmanial activity of these compounds are associated with their effects on the parasite plasma membrane, potentially leading to ionic leakage, subsequent disruption of mitochondrial membrane potential, and enhanced ROS generation.
{"title":"Antileishmanial, cytotoxic activities, and membrane rigidity effects of three synthetic compounds","authors":"Éder Jéferson Souza Cardoso , Ellyêssa Nascimento Borges , Kleber Santiago Freitas e Silva , Rodrigo Saar Gomes , Fabiana Taniguchi Boni , Jhon Kennedy Alves Pereira , Paula Maria Alexandre Mangoba , Luciano Morais Lião , Ricardo Menegatti , Antonio Alonso","doi":"10.1016/j.bbamem.2025.184450","DOIUrl":"10.1016/j.bbamem.2025.184450","url":null,"abstract":"<div><div>Three antileishmanial compounds incorporating a butylated hydroxytoluene (BHT) moiety and an acrylate-based Michael acceptor scaffold were rationally designed from the lead structures LQFM064 and LQFM332, which feature a chalcone-derived core. Their activities against <em>Leishmania</em> (<em>L</em>.) <em>amazonensis</em> were evaluated. Acrylate derivatives (<strong>5</strong>), (<strong>6</strong>), and (<strong>7</strong>) displayed IC<sub>50</sub> values comparable to miltefosine, while showing markedly lower cytotoxicity toward J774.A1 and differentiated THP-1 macrophages, along with reduced hemolytic potential. Spin-label electron paramagnetic resonance (EPR) spectroscopy revealed that treatment with these compounds induces membrane rigidity after 24 h in a concentration-dependent manner. This effect is unlikely due to direct membrane interaction, as it does not occur after short incubations or at low concentrations, suggesting a correlation with oxidative stress, such as lipid peroxidation and/or protein oxidation, likely triggered by elevated reactive oxygen species (ROS) production. In contrast, no oxidative stress-induced membrane rigidity was detected in uninfected macrophages, suggesting that nitric oxide production may mitigate oxidative damage in these cells. However, significant membrane rigidity was observed in <em>Leishmania</em>-infected macrophages at concentrations slightly above the IC<sub>50</sub> for amastigotes, indicating that the compounds may selectively target infected macrophages. Additionally, compound (<strong>5</strong>) exhibited moderate cytotoxicity in the rapidly proliferating J774.A1 macrophage line but displayed very low cytotoxicity in differentiated, non-proliferative THP-1 macrophages. Overall, this study suggests that the primary mechanisms underlying the antileishmanial activity of these compounds are associated with their effects on the parasite plasma membrane, potentially leading to ionic leakage, subsequent disruption of mitochondrial membrane potential, and enhanced ROS generation.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184450"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145010033","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01Epub Date: 2025-09-08DOI: 10.1016/j.bbamem.2025.184455
Leilismara Sousa Nogueira , Bárbara Martins Cordeiro , Gilvânia Aparecida Rabelo Cordeiro , Ana Carolina Pacheco Ramos , Douglas Donizetti Raimundo , Grazielle Aparecida Silva Maia , Cristiane de Oliveira Renó , Leonardo Nunes Medeiros , Luciana Estéfani Drumont Carvalho , Vanessa Faria Cortes , Hérica de Lima Santos , Carlos Frederico Leite Fontes , Leandro Augusto Barbosa
Lactoferrin (Lf) is an iron-binding glycoprotein involved in various biological functions, including iron metabolism and immune response. Bovine lactoferrin (bLf) has gained attention due to its potential therapeutic applications. This study investigates the effects of bLf on human erythrocyte membranes, focusing on Na,K-ATPase (NKA) modulation. Whole blood and erythrocyte ghost membranes were treated with different concentrations of bLf, and multiple assays were performed to assess enzyme activity, lipid peroxidation, and lipid membrane composition. First, we demonstrate that Fe+2 is not able to increase the activity of the NKA. Bovine lactoferrin, on the other hand, significantly increases NKA activity, particularly affecting the α3 isoform. Additionally, bLf treatment led to an increase in total phospholipids and cholesterol content, suggesting modifications in the erythrocyte membrane. Molecular docking analysis revealed a strong interaction between bLf and α3-NKA, but not α1 isoform, supporting a potential regulatory role. Furthermore, bLf reduced Fe3+ and Thiobarbituric Acid Reactive Substances (TBARS) levels, indicating a protective effect against oxidative stress. These findings suggest that bLf modulates erythrocyte membrane properties, and could be specific to α3-NKA activation, reinforcing the role of Fe+3 in modulating the NKA activity.
乳铁蛋白（Lf）是一种铁结合糖蛋白，参与多种生物功能，包括铁代谢和免疫反应。牛乳铁蛋白（bLf）因其潜在的治疗应用而备受关注。本研究探讨了bLf对人红细胞膜的影响，重点是Na， k - atp酶（NKA）的调节。用不同浓度的bLf处理全血和红细胞鬼膜，并进行多项测定，以评估酶活性、脂质过氧化和脂质膜组成。首先，我们证明了Fe+2不能增加NKA的活性。另一方面，牛乳铁蛋白显著增加NKA活性，特别是影响α3亚型。此外，bLf治疗导致总磷脂和胆固醇含量增加，表明红细胞膜发生了改变。分子对接分析显示，bLf与α3-NKA之间存在较强的相互作用，但与α1亚型不存在相互作用，支持其潜在的调控作用。此外，bLf降低了Fe3+和硫代巴比妥酸反应物质（TBARS）水平，表明对氧化应激具有保护作用。这些结果表明，bLf可以调节红细胞膜性质，并且可能特异性地激活α3-NKA，从而增强了Fe+3在调节NKA活性中的作用。
{"title":"Evaluation of the effects of bovine lactoferrin on the membrane of human erythrocytes","authors":"Leilismara Sousa Nogueira , Bárbara Martins Cordeiro , Gilvânia Aparecida Rabelo Cordeiro , Ana Carolina Pacheco Ramos , Douglas Donizetti Raimundo , Grazielle Aparecida Silva Maia , Cristiane de Oliveira Renó , Leonardo Nunes Medeiros , Luciana Estéfani Drumont Carvalho , Vanessa Faria Cortes , Hérica de Lima Santos , Carlos Frederico Leite Fontes , Leandro Augusto Barbosa","doi":"10.1016/j.bbamem.2025.184455","DOIUrl":"10.1016/j.bbamem.2025.184455","url":null,"abstract":"<div><div>Lactoferrin (Lf) is an iron-binding glycoprotein involved in various biological functions, including iron metabolism and immune response. Bovine lactoferrin (bLf) has gained attention due to its potential therapeutic applications. This study investigates the effects of bLf on human erythrocyte membranes, focusing on Na,K-ATPase (NKA) modulation. Whole blood and erythrocyte ghost membranes were treated with different concentrations of bLf, and multiple assays were performed to assess enzyme activity, lipid peroxidation, and lipid membrane composition. First, we demonstrate that Fe<sup>+2</sup> is not able to increase the activity of the NKA. Bovine lactoferrin, on the other hand, significantly increases NKA activity, particularly affecting the α3 isoform. Additionally, bLf treatment led to an increase in total phospholipids and cholesterol content, suggesting modifications in the erythrocyte membrane. Molecular docking analysis revealed a strong interaction between bLf and α3-NKA, but not α1 isoform, supporting a potential regulatory role. Furthermore, bLf reduced Fe<sup>3+</sup> and Thiobarbituric Acid Reactive Substances (TBARS) levels, indicating a protective effect against oxidative stress. These findings suggest that bLf modulates erythrocyte membrane properties, and could be specific to α3-NKA activation, reinforcing the role of Fe<sup>+3</sup> in modulating the NKA activity.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 8","pages":"Article 184455"},"PeriodicalIF":2.5,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145032579","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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