Topology surveillance of the lanosterol demethylase CYP51A1 by signal peptide peptidase.

Cleavage of transmembrane segments on target proteins by the aspartyl intramembrane protease signal peptide peptidase (SPP, encoded by HM13) has been linked to immunity, viral infection and protein quality control. How SPP recognizes its various substrates and specifies their fate remains elusive. Here, we identify the lanosterol demethylase CYP51A1 as an SPP substrate and show that SPP-catalysed cleavage triggers CYP51A1 clearance by endoplasmic reticulum-associated degradation (ERAD).

The Role of Cholesterol in M2 Clustering and Viral Budding Explained.

The influenza A M2 homotetrameric channel consists of four transmembrane (TM) and four amphipathic helices (AHs). This viral proton channel is suggested to form clusters in the catenoid budding neck areas in raft-like domains of the plasma membrane, resulting in cell membrane scission and viral release. The channel clustering environment is rich in cholesterol.

Difluorinated thromboxane A reveals crosstalk between platelet activatory and inhibitory pathways by targeting both the TP and IP receptors.

Background And Purpose: Thromboxane A (TXA) is a prostanoid produced during platelet activaton, important in enhancing platelet reactivity by activation of TP receptors. However, due to the short half-life, studying TXA signalling is challenging. To enhance our understanding of TP receptor-mediated platelet biology, we therefore synthesised mono and difluorinated TXA analogues and explored their pharmacology on heterologous and endogenously expressed TP receptor function.

Synthesis of lipid-linked precursors of the bacterial cell wall is governed by a feedback control mechanism in Pseudomonas aeruginosa.

Many bacterial surface glycans such as the peptidoglycan (PG) cell wall are built from monomeric units linked to a polyprenyl lipid carrier. How this limiting carrier is distributed among competing pathways has remained unclear. Here we describe the isolation of hyperactive variants of Pseudomonas aeruginosa MraY, the enzyme that forms the first lipid-linked PG precursor.

Biophysical basis of filamentous phage tactoid-mediated antibiotic tolerance in P. aeruginosa.

Inoviruses are filamentous phages infecting numerous prokaryotic phyla. Inoviruses can self-assemble into mesoscale structures with liquid-crystalline order, termed tactoids, which protect bacterial cells in Pseudomonas aeruginosa biofilms from antibiotics. Here, we investigate the structural, biophysical, and protective properties of tactoids formed by the P.

LipIDens: simulation assisted interpretation of lipid densities in cryo-EM structures of membrane proteins.

Cryo-electron microscopy (cryo-EM) enables the determination of membrane protein structures in native-like environments. Characterising how membrane proteins interact with the surrounding membrane lipid environment is assisted by resolution of lipid-like densities visible in cryo-EM maps. Nevertheless, establishing the molecular identity of putative lipid and/or detergent densities remains challenging.

LptM promotes oxidative maturation of the lipopolysaccharide translocon by substrate binding mimicry.

Insertion of lipopolysaccharide (LPS) into the bacterial outer membrane (OM) is mediated by a druggable OM translocon consisting of a β-barrel membrane protein, LptD, and a lipoprotein, LptE. The β-barrel assembly machinery (BAM) assembles LptD together with LptE at the OM. In the enterobacterium Escherichia coli, formation of two native disulfide bonds in LptD controls translocon activation.

The energetics and ion coupling of cholesterol transport through Patched1.

Patched1 (PTCH1) is a tumor suppressor protein of the mammalian Hedgehog (HH) signaling pathway, implicated in embryogenesis and tissue homeostasis. PTCH1 inhibits the G protein-coupled receptor Smoothened (SMO) via a debated mechanism involving modulating ciliary cholesterol accessibility. Using extensive molecular dynamics simulations and free energy calculations to evaluate cholesterol transport through PTCH1, we find an energetic barrier of ~15 to 20 kilojoule per mole for cholesterol export.

A feedback control mechanism governs the synthesis of lipid-linked precursors of the bacterial cell wall.

Many bacterial surface glycans such as the peptidoglycan (PG) cell wall, O-antigens, and capsules are built from monomeric units linked to a polyprenyl lipid carrier. How this limiting lipid carrier is effectively distributed among competing pathways has remained unclear for some time. Here, we describe the isolation and characterization of hyperactive variants of MraY, the essential and conserved enzyme catalyzing the formation of the first lipid-linked PG precursor called lipid I.

Cyclic di-AMP traps proton-coupled K transporters of the KUP family in an inward-occluded conformation.

Cyclic di-AMP is the only known essential second messenger in bacteria and archaea, regulating different proteins indispensable for numerous physiological processes. In particular, it controls various potassium and osmolyte transporters involved in osmoregulation. In Bacillus subtilis, the K/H symporter KimA of the KUP family is inactivated by c-di-AMP.

The Energetics and Ion Coupling of Cholesterol Transport Through Patched1.

Patched1 (PTCH1) is the principal tumour suppressor protein of the mammalian Hedgehog (HH) signalling pathway, implicated in embryogenesis and tissue homeostasis. PTCH1 inhibits the Class F G protein-coupled receptor Smoothened (SMO) via a debated mechanism involving modulating accessible cholesterol levels within ciliary membranes. Using extensive molecular dynamics (MD) simulations and free energy calculations to evaluate cholesterol transport through PTCH1, we find an energetic barrier of ~15-20 kJ mol for cholesterol export.

Supramolecular organization and dynamics of mannosylated phosphatidylinositol lipids in the mycobacterial plasma membrane.

() is the causative agent of tuberculosis (TB), a disease that claims ~1.6 million lives annually. The current treatment regime is long and expensive, and missed doses contribute to drug resistance.

Comparative Study of Receptor-, Receptor State-, and Membrane-Dependent Cholesterol Binding Sites in A and A Adenosine Receptors Using Coarse-Grained Molecular Dynamics Simulations.

We used coarse-grained molecular dynamics (CG MD) simulations to study protein-cholesterol interactions for different activation states of the A adenosine receptor (AR) and the A adenosine receptor (AR) and predict new cholesterol binding sites indicating amino acid residues with a high residence time in three biologically relevant membranes. Compared to 1-palmitoyl-2-oleoyl--glycero-3-phosphocholine (POPC)-cholesterol and POPC-phosphatidylinositol-bisphosphate (PIP)-cholesterol, the plasma mimetic membrane best described the cholesterol binding sites previously detected for the inactive state of AR and revealed the binding sites with long-lasting amino acid residues. We observed that using the plasma mimetic membrane and plotting residues with cholesterol residence time ≥2 μs, our CG MD simulations captured most obviously the cholesterol-protein interactions.

Cardiolipin, and not monolysocardiolipin, preferentially binds to the interface of complexes III and IV.

The mitochondrial electron transport chain comprises a series of protein complexes embedded in the inner mitochondrial membrane that generate a proton motive force oxidative phosphorylation, ultimately generating ATP. These protein complexes can oligomerize to form larger structures called supercomplexes. Cardiolipin (CL), a conical lipid, unique within eukaryotes to the inner mitochondrial membrane, has proven essential in maintaining the stability and function of supercomplexes.

Inhibited KdpFABC transitions into an E1 off-cycle state.

KdpFABC is a high-affinity prokaryotic K uptake system that forms a functional chimera between a channel-like subunit (KdpA) and a P-type ATPase (KdpB). At high K levels, KdpFABC needs to be inhibited to prevent excessive K accumulation to the point of toxicity. This is achieved by a phosphorylation of the serine residue in the TGES motif in the A domain of the pump subunit KdpB (KdpB).

Seipin concentrates distinct neutral lipids via interactions with their acyl chain carboxyl esters.

Lipid droplets (LDs) are essential for cellular lipid homeostasis by storing diverse neutral lipids (NLs), such as triacylglycerol (TAG), steryl esters (SE), and retinyl esters (RE). A proper assembly of TAG-containing LDs at the ER requires Seipin, a conserved protein often mutated in lipodystrophies. Here, we show that the yeast Seipin Sei1 and its partner Ldb16 also promote the storage of other NL in LDs.

Interaction With the Lipid Membrane Influences Fentanyl Pharmacology.

Overdose deaths from fentanyl have reached epidemic proportions in the USA and are increasing worldwide. Fentanyl is a potent opioid agonist that is less well reversed by naloxone than morphine. Due to fentanyl's high lipophilicity and elongated structure we hypothesised that its unusual pharmacology may be explained by its interactions with the lipid membrane on route to binding to the μ-opioid receptor (MOPr).

The guidance and adhesion protein FLRT2 dimerizes in cis via dual small-X-small transmembrane motifs.

Fibronectin Leucine-rich Repeat Transmembrane (FLRT 1-3) proteins are a family of broadly expressed single-spanning transmembrane receptors that play key roles in development. Their extracellular domains mediate homotypic cell-cell adhesion and heterotypic protein interactions with other receptors to regulate cell adhesion and guidance. These in trans FLRT interactions determine the formation of signaling complexes of varying complexity and function.

Rate-limiting transport of positively charged arginine residues through the Sec-machinery is integral to the mechanism of protein secretion.

Transport of proteins across and into membranes is a fundamental biological process with the vast majority being conducted by the ubiquitous Sec machinery. In bacteria, this is usually achieved when the SecY-complex engages the cytosolic ATPase SecA (secretion) or translating ribosomes (insertion). Great strides have been made towards understanding the mechanism of protein translocation.

Peptidoglycan biosynthesis is driven by lipid transfer along enzyme-substrate affinity gradients.

Maintenance of bacterial cell shape and resistance to osmotic stress by the peptidoglycan (PG) renders PG biosynthetic enzymes and precursors attractive targets for combating bacterial infections. Here, by applying native mass spectrometry, we elucidate the effects of lipid substrates on the PG membrane enzymes MraY, MurG, and MurJ. We show that dimerization of MraY is coupled with binding of the carrier lipid substrate undecaprenyl phosphate (C-P).

Structure, substrate recognition and initiation of hyaluronan synthase.

Hyaluronan is an acidic heteropolysaccharide comprising alternating N-acetylglucosamine and glucuronic acid sugars that is ubiquitously expressed in the vertebrate extracellular matrix. The high-molecular-mass polymer modulates essential physiological processes in health and disease, including cell differentiation, tissue homeostasis and angiogenesis. Hyaluronan is synthesized by a membrane-embedded processive glycosyltransferase, hyaluronan synthase (HAS), which catalyses the synthesis and membrane translocation of hyaluronan from uridine diphosphate-activated precursors.

Specific interactions of peripheral membrane proteins with lipids: what can molecular simulations show us?

Peripheral membrane proteins (PMPs) can reversibly and specifically bind to biological membranes to carry out functions such as cell signalling, enzymatic activity, or membrane remodelling. Structures of these proteins and of their lipid-binding domains are typically solved in a soluble form, sometimes with a lipid or lipid headgroup at the binding site. To provide a detailed molecular view of PMP interactions with the membrane, computational methods such as molecular dynamics (MD) simulations can be applied.

PyLipID: A Python Package for Analysis of Protein-Lipid Interactions from Molecular Dynamics Simulations.

Lipids play important modulatory and structural roles for membrane proteins. Molecular dynamics simulations are frequently used to provide insights into the nature of these protein-lipid interactions. Systematic comparative analysis requires tools that provide algorithms for objective assessment of such interactions.

Structure of the hexameric fungal plasma membrane proton pump in its autoinhibited state.

The fungal plasma membrane H-ATPase Pma1 is a vital enzyme, generating a proton-motive force that drives the import of essential nutrients. Autoinhibited Pma1 hexamers in the plasma membrane of starving fungi are activated by glucose signaling and subsequent phosphorylation of the autoinhibitory domain. As related P-type adenosine triphosphatases (ATPases) are not known to oligomerize, the physiological relevance of Pma1 hexamers remained unknown.