Molecular Membrane Biology最新文献

Membrane-bound pyrophosphatases (PPases) are involved in the adaption of organisms to stress conditions, which was substantiated by numerous plant transgenic studies with H⁺-PPase yet devoid of any correlated evidences for other two subfamilies, Na⁺-PPase and Na⁺,H⁺-PPase. Herein, we demonstrate the gene cloning and functional evaluation of the membrane-bound PPase (CmPP) of the human gut microbe Clostridium methylpentosum. The CmPP gene encodes a single polypeptide of 699 amino acids that was predicted as a multi-spanning membrane and K⁺-dependent Na⁺,H⁺-PPase. Heterologous expression of CmPP could significantly enhance the salt tolerance of both Escherichia coli and Saccharomyces cerevisiae, and this effect in yeast could be fortified by N-terminal addition of a vacuole-targeting signal peptide from the H⁺-PPase of Trypanosoma cruzi. Furthermore, introduction of CmPP could remarkably improve the salt tolerance of tobacco, implying its potential use in constructing salt-resistant transgenic crops. Consequently, the possible mechanisms of CmPP to underlie salt tolerance are discussed.

A molecular dynamics (MD) simulation with atomistic details was performed to examine the partitioning and transport behavior of moderately cytotoxic ionic liquids (ILs), namely choline bis(2-ethylhexyl) phosphate (CBEH), choline bis(2,4,4-trimethylpentyl) phosphinate (CTMP) and choline O,O-diethyl dithiophosphate (CDEP) in a fully hydrated dipalmitoylphosphatidylcholine (DPPC) bilayer in the fluid phase at 323 K. The structure of ILs was so selected to understand if the role of dipole and dispersion forces in the ILs distribution in the membrane can be possible. Several analyses including mass density, electrostatic potential, order parameter, diffusion coefficients and hydrogen bond formation, was carried out to determine the precise location of the anionic species inside the membrane. Moreover, the potential of the mean force (PMF) method was used to calculate free energy profile for transferring anionic species from the DPPC membrane into the bulk water. While less cytotoxic DEP is located within the bulk water, more cytotoxic TMP and BEH ILs were found to remain in the membrane and the energy barrier for crossing through the bilayer center of BEH was higher. Various ILs have no significant effect on P-N vector. The thickness of lipid bilayer decreased in all systems comprising ILs, while area per lipid increased.

Membrane proteins are ubiquitous in biology and are key targets for therapeutic development. Despite this, our structural understanding has lagged behind that of their soluble counterparts. This review provides an overview of this important field, focusing in particular on the recent resurgence of electron microscopy (EM) and the increasing role it has to play in the structural studies of membrane proteins, and illustrating this through several case studies. In addition, we examine some of the challenges remaining in structural determination, and what steps are underway to enhance our knowledge of these enigmatic proteins.

Geranylgeranyl diphosphate is a 20-carbon isoprenoid phospholipid whose lipid moiety can be post-translationally incorporated into proteins to promote membrane association. The process of geranylgeranylation has been implicated in anti-proliferative effects of clinical agents that inhibit enzymes of the mevalonate pathway (i.e. statins and nitrogenous bisphosphonates) as well as experimental agents that deplete geranylgeranyl diphosphate. Inhibitors of geranylgeranyl diphosphate synthase are an attractive way to block geranylgeranylation because they possess a calcium-chelating substructure to allow localization to bone and take advantage of a unique position of the enzyme within the biosynthetic pathway. Here, we describe recent advances in geranylgeranyl diphosphate synthase expression and inhibitor development with a particular focus on the molecular mechanisms that link geranylgeranyl diphosphate to cell proliferation via geranylgeranylated small GTPases.

To investigate the effect of fluorescent probe on the properties of membranes, we studied model membranes composed of 1,2- dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1-palmitoyl 2-oleoyl-sn-glycero-3-phosphocholine (POPC) in the presence and absence of fluorescent probe. The morphology of giant unilamellar vesicles (GUVs) has been observed as a function of temperature and composition by fluorescence microscopy using NBD-DOPE or C₆-NBD-PC as the probe. The phase behavior of model membranes containing no fluorescent probe was investigated by ²H-NMR spectroscopy. We found that the bright phase observed on GUVs was the fluid phase enriched in POPC and the dark phase was the gel phase enriched in DPPC. NBD-DOPE and C₆-NBD-PC preferentially participated in the fluid-phase domains when GUVs were in the gel + fluid phase coexistence. Inclusion of both fluorescent probes (1 mol%) lowered the transition temperature of POPC/DPPC membranes. In addition, C₆-NBD-PC exhibited a stronger effect than NBD-DOPE, which was considered to be associated with the structures of fluorescent molecules.

The PapC usher, a β-barrel pore in the outer membrane of uropathogenic Escherichia coli, is used for assembly of the P pilus, a key virulence factor in bacterial colonization of human kidney cells. Each PapC protein is composed of a 24-stranded β-barrel channel, flanked by N- and C-terminal globular domains protruding into the periplasm, and occluded by a plug domain (PD). The PD is displaced from the channel towards the periplasm during pilus biogenesis, but the molecular mechanism for PD displacement remains unclear. Two structural features within the β-barrel, an α-helix and β5-6 hairpin loop, may play roles in controlling plug stabilization. Here we have tested clusters of residues at the interface of the plug, barrel, α-helix and hairpin, which participate in electrostatic networks. To assess the roles of these residues in plug stabilization, we used patch-clamp electrophysiology to compare the activity of wild-type and mutant PapC channels containing alanine substitutions at these sites. Mutations interrupting each of two salt bridge networks were relatively ineffective in disrupting plug stabilization. However, mutation of two pairs of arginines located at the inner and the outer surfaces of the PD resulted in an enhanced propensity for plug displacement. One arginine pair involved in a repulsive interaction between the linkers that tether the plug to the β-barrel was particularly sensitive to mutation. These results suggest that plug displacement, which is necessary for pilus assembly and translocation, may require a weakening of key electrostatic interactions between the plug linkers, and the plug and the α-helix.

Modification of proteins with the lipid palmitate, a process called palmitoylation, is important for the normal function of neuronal cells. However, most attention has focused on how palmitoylation regulates the targeting and trafficking of neurotransmitter receptors and non-enzymatic scaffold proteins. In this review we discuss recent studies that suggest that palmitoylation also plays additional roles in neurons by controlling the localization, interactions and perhaps even the activity of protein kinases that play key roles in physiological neuronal regulation and in neuropathological processes.