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[en] The investigation of the interaction of peptides with cell membranes is the focus of active research. It can enhance the understanding of basic membrane functions such as membrane transport, fusion, and signaling processes, and it may shed light on potential applications of peptides in biomedicine. In this review, we will present current advances in computational studies on the interaction of different types of peptides with the cell membrane. Depending on the properties of the peptide, membrane, and external environment, the peptide–membrane interaction shows a variety of different forms. Here, on the basis of recent computational progress, we will discuss how different peptides could initiate membrane pores, translocate across the membrane, induce membrane endocytosis, produce membrane curvature, form fibrils on the membrane surface, as well as interact with functional membrane proteins. Finally, we will present a conclusion summarizing recent progress and providing some specific insights into future developments in this field. (topical review)
[en] Substrate transport through the membrane protein maltoporin is facilitated by an affinity site in the channel. The analysis of the ion current fluctuations induced by penetration of the sugar into the channel yields the kinetic constants. Modification of the affinity site by replacing the aromatic residues suggests that nature has optimized the channel protein for maximum affinity at the extracellular side, as well as for an increased off-rate to eject a sugar trapped in the pore towards the periplasmic side
[en] The transport of biopolymers through large membrane channels is a ubiquitous process in biology. It is central to processes such as gene transfer by transduction and RNA transport through nuclear pore complexes. The transport of polymers through nanoscopic channels is also of interest to physicists and chemists studying the effects of steric, hydrodynamic, and electrostatic interactions between polymers and confining walls. Single-channel ion current measurements have been recently used to study the transport of biopolymers, and in particular single-stranded DNA and RNA molecules, through nanometre-size channels. Under the influence of an electric field, the negatively charged polynucleotides can be captured and drawn through the channel in a process termed 'translocation'. During translocation, the ion current flowing through the channel is mostly blocked, indicating the presence of the polymer inside the channel. The current blockades were found to be sensitive to the properties of the biopolymers such as their nucleotide composition, length, and secondary structure, and to physical parameters such as the driving field intensity, temperature, and ionic strength. These blockades are therefore a rich source of information regarding the dynamics of polynucleotides in the pore. The translocation process is separated into its two main steps: (a) polymer 'capture' in which one of the polymer's ends is threaded a small distance through the channel, and (b) polymer sliding through the channel. The experimental and theoretical efforts to elucidate polymer capture and the transport dynamics of biopolymers in nanoscopic pores are reviewed in this article. (topical review)
[en] The Gram-negative bacterial cell envelope is a complex structure. It consists of a cytoplasmic inner membrane, the periplasm and an outer membrane. The outer membrane composes of a phospholipid inner leaflet and an outer leaflet of lipopolysaccharide (LPS) as well as integral membrane proteins. The challenge in studying the structures and functions at the outer membrane is in creating simple, but yet representative models of the membrane. Current models consist of solid- supported bilayers containing the lipid and protein components of the bacterial outer membrane. Although this makes for a good representation of a bilayer the fluidity of the lipid layers are not truly represented. One method to mimic membrane fluidity is to create lipid monolayers on water surfaces. This is relatively straight forward to achieve using simple amphipathic phospholipids. However, creating such layers that truly represent the bacterial cell surface is more difficult due to LPS being a large water soluble molecule. Here we describe how creating monolayers using LPS were achieved. The LPS monolayers were found to be stable as shown by reproducible pressure-area isotherms. The structure of the LPS monolayers were characterised using neutron and X-ray reflectometry and grazing incidence X-ray diffraction. To add further complexity the LPS was mixed with the E. coli porin, OmpF. Thus mixed monolayers of OmpF and LPS were created making a good representation of the Gram-negative bacterial cell surface. Deuterated versions of both the OmpF and LPS were produced and by using different combinations of deuterated and hydrogenated material a detailed picture of the monolayer is described through neutron reflectometry.
[en] Functional Channelrhodopsin-2 (ChR2) overexpression of about 104 channels/μm2 in the plasma membrane of HEK293 cells was studied by patch-clamp and freeze-fracture electron microscopy. Simultaneous electrorotation measurements revealed that ChR2 expression was accompanied by a marked increase of the area-specific membrane capacitance (Cm). The Cm increase apparently resulted partly from an enlargement of the size and/or number of microvilli. This is suggested by a relatively large Cm of 1.15 ± 0.08 μF/cm2 in ChR2-expressing cells measured under isotonic conditions. This value was much higher than that of the control HEK293 cells (0.79 ± 0.02 μF/cm2). However, even after complete loss of microvilli under strong hypoosmolar conditions (100 mOsm), the ChR2-expressing cells still exhibited a significantly larger Cm (0.85 ± 0.07 μF/cm2) as compared to non-expressing control cells (0.70 ± 0.03 μF/cm2). Therefore, a second mechanism of capacitance increase may involve changes in the membrane permittivity and/or thickness due to the embedded ChR2 proteins
[en] Background and Aims: Sterols have been reported to modulate conformation and hence the function of several membrane proteins. One such group is the Chloride Intracellular Ion Channel (CLIC) family of proteins. The CLIC protein family consists of six evolutionarily conserved protein members in vertebrates. These proteins are unusual, existing as both monomeric soluble proteins and as membrane bound proteins. We now for the first time demonstrate that the spontaneous membrane insertion of CLIC1 is dependent on the presence of cholesterol in membranes. Our novel findings also extend to the identification of a cholesterol-binding domain within CLIC1 that facilitates the spontaneous membrane insertion of the protein into membranes containing cholesterol. Methods: CLIC1 wild type (WT) and mutant proteins were purified by Ni-NTA followed by size‐exclusion chromatography. Langmuir monolayer film balance experiments were carried out using 1-Palmitoyl-2-oleoylphosphatidylcholine (POPC) alone, or in a 5:1 mole ratio combination with either one of the following sterols: Cholesterol (CHOL), β-Sitosterol (SITO), Ergosterol (ERG), Hydroxyecdysone (HYD) or Cholestane (CHOS). WT CLIC1 or mutant versions of CLIC1 were then injected into the aqueous subphase under the lipid film. Results: In lipid monolayers lacking sterols, CLIC1 did not insert. However significant membrane insertion occurred when CLIC1 was added to membranes containing cholesterol. Substitution of membrane cholesterol with either HYD, SITO or ERG, not only increased CLIC1’s membrane interaction but also increased its rate of insertion. Conversely, CLIC1 showed no insertion into monolayers containing CHOS, which lacked the intact sterol 3β-OH group. CLIC1 mutants G18A and G22A, did not insert in POPC:CHOL monolayers whereas the C24A mutant showed membrane insertion equivalent to WT CLIC1. X-ray and Neutron reflectivity, along with Small Angle X-ray Scattering techniques were subsequently used to probe structural features of the membrane bound CLIC1 and CLIC1-Cholesterol complex in solution. Conclusion: These findings confirm that the GXXXG motif within CLIC1 acts as a sterol binding site facilitating the protein’s membrane interaction and insertion. Critical to this process of spontaneous membrane insertion is the presence of an intact 3β-OH group within the sterol structure itself. Furthermore, additional double bonds and methylation of the steroid skeleton enhanced CLIC1 membrane insertion.
[en] Flexoelectricity provides a reciprocal relationship between electricity and mechanics in membranes, i.e., between membrane curvature and polarization. Experimental evidence of biomembrane flexoelectricity (including direct and converse flexoelectric effect) is reviewed. Biological implications of flexoelectricity in membrane transport, membrane contact, mechanosensitivity, electromotility and hearing are underlined. Flexoelectricity enables membrane structures to function like soft micro- and nano-machines, sensors and actuators, thus providing important input to molecular electronics applications
[en] Varicella-zoster virus (VZV) encodes two multiply inserted membrane proteins, open reading frame (ORF) 39 protein (ORF39p) and glycoprotein K (gK). The HSV-1 homologs of these proteins are believed to act in conjunction with each other during viral egress and cell-cell fusion, and they directly influence each other's intracellular trafficking. However, ORF39p and VZV gK have received very limited study largely due to difficulties in producing antibodies to these highly hydrophobic proteins. To overcome this obstacle, we introduced epitope tags into both ORF39p and gK and examined their intracellular distributions in transfected and infected cells. Our data demonstrate that both ORF39p and gK accumulate predominately in the ER of cultured cells when expressed in the absence of other VZV proteins or when coexpressed in isolation from other VZV proteins. Therefore, the transport of VZV ORF39p and gK does not exhibit the functional interdependence seen in their HSV-1 homologs. However, during infection, the primary distributions of ORF39p and gK shift from the ER to the Golgi, and they are also found in the plasma membrane indicating that their intracellular trafficking during infection depends on other VZV-encoded proteins. During infection, ORF39p and gK tightly colocalize with VZV envelope glycoproteins B, E and H; however, the coexpression of ORF39p or gK with other individual viral glycoproteins is insufficient to alter the transport of either ORF39p or gK
[en] Dodecyl-β-d-selenomaltoside in a leukotriene C4 synthase crystal exhibited sufficient anomalous diffraction for multiwavelength anomalous diffraction phasing. Dodecyl-β-d-selenomaltoside (SeDDM) is a seleno-detergent with a β-glycosidic seleno-ether in place of the ether moiety in dodecyl-β-d-maltoside. Seleno-detergents are candidates for heavy-atom agents in experimental phasing of membrane proteins in protein crystallography. Crystals of a nuclear membrane-embedded enzyme, leukotriene C4 synthase (LTC4S), in complex with SeDDM were prepared and a multiwavelength anomalous diffraction (MAD) experiment was performed. The SeDDM in the LTC4S crystal exhibited sufficient anomalous diffraction for determination of the structure using MAD phasing