Journal of Membrane Biology最新文献

Many pathogenic bacteria utilize their complicated appalling arsenal, bacterial virulence factors, to attack host cells by damaging the host cell membrane and neutralizing host defense mechanisms. Bacterial pore-forming proteins (PFPs) are one of them, they include a distinct class of secreted soluble toxin monomers, which binds to the specific cell surface receptors and /or lipids, oligomerizes as an amphipathic transmembrane pore complex on host cell membranes, and deforms the integrity of the plasma membrane. Researchers have focused on characterizing the structure and function of different Pore Forming Toxins (PFTs) from various organisms, where most of the structural studies employed X-ray crystallography, single-particle cryo-EM, and cryo-electron tomography. However, historically, most of these previous studies focused on using detergent to solubilize and oligomerize the PFTs. Additionally, previous studies have also shown that lipid membranes and lipid components, including cell surface receptors, play a critical role in pore formation and oligomerization. However, there are limited studies available that aim to resolve the structure and function of PFTs in liposomes. In this review article, we majorly focused on structural and functional studies of pore-forming toxins in the presence of detergents, lipid nanodiscs, and liposomes. We will also discuss the challenges and benefits of using liposomes to study pore-forming proteins in more biologically relevant membrane environments.

The complement pathway is one of the most ancient elements of the host's innate response and includes a set of protein effectors that rapidly react against pathogens. The late stages of the complement reaction are broadly categorised into two major outcomes. Firstly, C5a receptors, expressed on membranes of host cells, are activated by C5a to generate pro-inflammatory responses. Secondly, target cells are lysed by a hetero-oligomeric pore known as the membrane attack complex (MAC) that punctures the cellular membrane, causing ion and osmotic flux. Generally, several membrane-bound and soluble inhibitors protect the host membrane from complement damage. This includes inhibitors against the MAC, such as clusterin and CD59. This review addresses the most recent molecular and structural insights behind the activation and modulation of the integral membrane proteins, the C5a receptors (C5aR1 and C5aR2), as well as the regulation of MAC assembly. The second aspect of the review focuses on the molecular basis behind inflammatory diseases that are reflective of failure to regulate the terminal complement effectors. Although each arm is unique in its function, both pathways may share similar outcomes in these diseases. As such, the review outlines potential synergy and crosstalk between C5a receptor activation and MAC-mediated cellular responses.

Pore-forming toxins (PFTs) belong to a class of proteins expressed by bacteria to initiate infections by unregulated pore formation on the plasma membrane of host cells. Although cholesterol is a key sterol motif that promotes toxin activity, the influence of oxysterols, upregulated in senescent cells or in other inflammatory disorders, on lytic activity has not received much attention. Using all-atom molecular dynamics simulations, we study the changes to the sterol binding landscape of membrane-inserted cytolysin A (ClyA), an $\alpha$ -PFT expressed by E. coli, in the presence of tail-oxidized 25-hydroxycholesterol (25-HC) in a palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC):cholesterol:25-HC (70:20:10) membrane. 25-HC was found to entirely replace previously identified cholesterol binding hotspots [PNAS,115 7323-7330] between the membrane-inserted $\beta$ -tongue motifs with binding lifetimes on the order of microseconds. Although the overall sterol occupancy is lower for the N-terminal helix motif that forms the lining of the water channel, 25-HC binding is less when compared with cholesterol. The presence of the additional OH group on the 25th carbon enhances interactions with polar residues of the $\beta$ -tongue, increasing 25-HC binding times by several fold when compared with cholesterol. We discuss the implications of this enhanced oxysterol interaction on pore formation of the $\alpha$ family of toxins such as ClyA, in contrast with the cholesterol-dependent cytolysins, where oxysterols have been shown to be detrimental to pore formation.

Protein nanopores are emerging as versatile single-molecule sensors with broad applications in DNA and protein sequencing. However, their narrow size restricts the range of detectable analytes, necessitating the development of advanced nanopores to broaden their applications in biotechnology. This review highlights a natural hetero-oligomeric porin, Nocardia farcinica porin AB (NfpAB), based on the Gram-positive mycolata, Nocardia farcinica. The pore comprises two subunits, NfpA and NfpB, that combine to form a stable structure with a unique pore geometry, asymmetrical shape, and charge distribution. Single-channel electrical recordings demonstrate that NfpAB forms stable, high-conductance channels suitable for sensing charged molecules, particularly cationic polypeptides and cyclic sugars. This pore offers advantages such as enhanced control over molecular interactions due to densely crowded charged residues, thus allowing the quantification of voltage-dependent translocation kinetics. Notably, NfpAB contains intrinsic cysteines in the pore lumen, providing an accessible site for thiol-based reactions and attachment of molecular adapters. We propose that such hetero-oligomeric pores will be effective for several applications in nanopore technology for biomolecular detection and sequencing.

Plants are attacked by various pathogens that secrete a variety of effectors to damage host cells and facilitate infection. One of the largest and so far understudied microbial protein families of effectors is necrosis- and ethylene-inducing peptide-1-like proteins (NLPs), which are involved in important plant diseases. Many NLPs act as cytolytic toxins that cause cell death and tissue necrosis by disrupting the plant's plasma membrane. Their mechanism of action is unique and leads to the formation of small, transient membrane ruptures. Here, we capture the interaction of the cytotoxic model NLP from the oomycete Pythium aphanidermatum, NLP_Pya, with plant cell-mimicking membranes of giant unilamellar vesicles (GUVs) and tobacco protoplasts using confocal fluorescence microscopy. We show that the permeabilization of GUVs by NLP_Pya is concentration- and time-dependent, confirm the small size of the pores by observing the inability of NLP_Pya monomers to pass through them, image the morphological changes of GUVs at higher concentrations of NLP_Pya and confirm its oligomerization on the membrane of GUVs. In addition, NLP_Pya bound to plasma membranes of protoplasts, which showed varying responses. Our results provide new insights into the interaction of NLP_Pya with model lipid membranes containing plant-derived sphingolipids.

In this study, we describe the effect of the noble gas, xenon on the electrical properties of tethered lipid bilayer membranes, (tBLMs), including the effect of xenon on the activation energy for electrical conduction through the tBLM. Such studies benefit from the stability of a tethered membrane given the wide range of temperatures that are scanned and the time required for these measurements. The results indicate that xenon increases the activation energy for electrical conduction through bilayers and decreases the average pore size that dominates the electrical conductance of the lipid bilayers at low voltages. Xenon possesses a high affinity for lipid membranes and is a potent general anaesthetic. Its anaesthetic potency is possibly associated with its effects on proteins embedded in the lipid membranes.

Ion channels play an integral role in the normal functioning of the brain. They regulate neuronal electrical properties like synaptic activity, generation of action potentials, maintenance of resting membrane potential and neuronal plasticity, and modulate the physiology of non-neuronal cells like astrocytes and microglia. Dysregulation of ionic homeostasis and channelopathies are associated with various neurological disorders, including Alzheimer's disease (AD). Several families of ion channels are associated with AD pathophysiology and progression. In this review, we outline the current research centered around ion channel dysregulation during AD and discuss briefly the possibility of using ion channels as therapeutic targets.

Antimicrobial peptides are part of the innate immune response and show their antimicrobial activity by forming pores, followed by disintegration of the membrane. Cholesterol in the membrane can affect the pore formation process, as cholesterol is known to alter the permeability and elastic properties of the membrane. The present research systematically explores the role of cholesterol in modulating the interaction of the antimicrobial peptide NK-2 with phospholipid membranes, as well as the processes of pore formation induced by NK-2 within the membrane. Large unilamellar vesicles (LUVs) and giant unilamellar vesicles (GUVs) made from DOPC-DOPG and Egg PC with varying cholesterol concentrations have been studied using a variety of experimental techniques. The present study revealed that both the magnitude of zeta potential and surface charge density diminished as cholesterol concentrations increased at an intermediate NK-2 concentration. The proliferation of the size distributions of LUVs containing cholesterol when exposed to NK-2 indicates the occurrence of vesicle aggregation. The phase contrast micrographs of GUVs as well as the calcein release experiments on LUVs show evidence of pores. Notably, the incorporation of cholesterol into the membrane was found to have a significant effect on both the permeability of the membrane and the kinetics of the pore formation process. This biophysical research contributes essential knowledge regarding the role of cholesterol in influencing the antimicrobial efficacy of the membrane.