Lipid interactions with bacterial channels: fluorescence studies

Powl, Andrew M.; Carney, Joanne; Marius, Phedra; East, J. Malcolm; Lee, A.G.

doi:10.1042/bst20050905

Cited by 14 publications

(7 citation statements)

References 18 publications

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“…The very special properties of tryptophan make it among the more informative amino acid replacements that can be made. 15 , 32 , 34 , 47 , 62 , 63 Care must be taken with membrane proteins because of the known role of this amino acid in controlling topology by anchoring the periphery of transmembrane helices to the phospholipid headgroups of the bilayer. 64 , 65 Mechanosensitive channels are notable in lacking these “anchorages” but do contain Trp at other positions.…”

Section: Discussionmentioning

confidence: 99%

Properties of the Mechanosensitive Channel MscS Pore Revealed by Tryptophan Scanning Mutagenesis

Rasmussen

Singh

et al. 2015

Biochemistry

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Bacterial mechanosensitive channels gate when the transmembrane turgor rises to levels that compromise the structural integrity of the cell wall. Gating creates a transient large diameter pore that allows hydrated solutes to pass from the cytoplasm at rates close to those of diffusion. In the closed conformation, the channel limits transmembrane solute movement, even that of protons. In the MscS crystal structure (Protein Data Bank entry 2oau), a narrow, hydrophobic opening is visible in the crystal structure, and it has been proposed that a vapor lock created by the hydrophobic seals, L105 and L109, is the barrier to water and ions. Tryptophan scanning mutagenesis has proven to be a highly valuable tool for the analysis of channel structure. Here Trp residues were introduced along the pore-forming TM3a helix and in selected other parts of the protein. Mutants were investigated for their expression, stability, and activity and as fluorescent probes of the physical properties along the length of the pore. Most Trp mutants were expressed at levels similar to that of the parent (MscS YFF) and were stable as heptamers in detergent in the presence and absence of urea. Fluorescence data suggest a long hydrophobic region with low accessibility to aqueous solvents, extending from L105/L109 to G90. Steady-state fluorescence anisotropy data are consistent with significant homo-Förster resonance energy transfer between tryptophan residues from different subunits within the narrow pore. The data provide new insights into MscS structure and gating.

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Section: Discussionmentioning

confidence: 99%

Properties of the Mechanosensitive Channel MscS Pore Revealed by Tryptophan Scanning Mutagenesis

Rasmussen

Singh

et al. 2015

Biochemistry

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“…Thus, long-chain acyl-CoA esters, a metabolically active form of free fatty acids, activate K ATP channels ( Larsson et al, 1996 ; Bränström et al, 1997 ). Further, phosphatidic acid interacts with a variety of proteins ( Kooijman and Burger, 2009 ) and activates, for instance, the bacterial mechanosensitive channel of large conductance (MscL; Powl et al, 2005a , b ). Lastly, even K V 7.2/K V 7.3 channels can be kept active by phospholipids other than PI(4,5)P 2 that have at least an acyl chain and a phosphate head group ( Telezhkin et al, 2012 ).…”

Section: Discussionmentioning

confidence: 99%

Regulation of voltage-gated potassium channels by PI(4,5)P2

Kruse

Hammond

Hille

2012

Journal of General Physiology

103

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Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) regulates activities of numerous ion channels including inwardly rectifying potassium (Kir) channels, KCNQ, TRP, and voltage-gated calcium channels. Several studies suggest that voltage-gated potassium (KV) channels might be regulated by PI(4,5)P2. Wide expression of KV channels in different cells suggests that such regulation could have broad physiological consequences. To study regulation of KV channels by PI(4,5)P2, we have coexpressed several of them in tsA-201 cells with a G protein–coupled receptor (M1R), a voltage-sensitive lipid 5-phosphatase (Dr-VSP), or an engineered fusion protein carrying both lipid 4-phosphatase and 5-phosphatase activity (pseudojanin). These tools deplete PI(4,5)P2 with application of muscarinic agonists, depolarization, or rapamycin, respectively. PI(4,5)P2 at the plasma membrane was monitored by Förster resonance energy transfer (FRET) from PH probes of PLCδ1 simultaneously with whole-cell recordings. Activation of Dr-VSP or recruitment of pseudojanin inhibited KV7.1, KV7.2/7.3, and Kir2.1 channel current by 90–95%. Activation of M1R inhibited KV7.2/7.3 current similarly. With these tools, we tested for potential PI(4,5)P2 regulation of activity of KV1.1/KVβ1.1, KV1.3, KV1.4, and KV1.5/KVβ1.3, KV2.1, KV3.4, KV4.2, KV4.3 (with different KChIPs and DPP6-s), and hERG/KCNE2. Interestingly, we found a substantial removal of inactivation for KV1.1/KVβ1.1 and KV3.4, resulting in up-regulation of current density upon activation of M1R but no changes in activity upon activating only VSP or pseudojanin. The other channels tested except possibly hERG showed no alteration in activity in any of the assays we used. In conclusion, a depletion of PI(4,5)P2 at the plasma membrane by enzymes does not seem to influence activity of most tested KV channels, whereas it does strongly inhibit members of the KV7 and Kir families.

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“…It has been shown that different lipids have different binding affinities on the surface of membrane proteins [74] and that the specific lipid-protein interactions probably play a role in regulating the activity and/or partitioning of certain proteins such as the yeast cytochrome bc 1 complex [75]. However, various evidence exists that generic interaction mechanisms in terms of for example elastic properties of the membrane are also important for a number of membrane proteins [47].…”

Section: Discussionmentioning

confidence: 99%

Assessing the Nature of Lipid Raft Membranes

et al. 2007

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The paradigm of biological membranes has recently gone through a major update. Instead of being fluid and homogeneous, recent studies suggest that membranes are characterized by transient domains with varying fluidity. In particular, a number of experimental studies have revealed the existence of highly ordered lateral domains rich in sphingomyelin and cholesterol (CHOL). These domains, called functional lipid rafts, have been suggested to take part in a variety of dynamic cellular processes such as membrane trafficking, signal transduction, and regulation of the activity of membrane proteins. However, despite the proposed importance of these domains, their properties, and even the precise nature of the lipid phases, have remained open issues mainly because the associated short time and length scales have posed a major challenge to experiments. In this work, we employ extensive atom-scale simulations to elucidate the properties of ternary raft mixtures with CHOL, palmitoylsphingomyelin (PSM), and palmitoyloleoylphosphatidylcholine. We simulate two bilayers of 1,024 lipids for 100 ns in the liquid-ordered phase and one system of the same size in the liquid-disordered phase. The studies provide evidence that the presence of PSM and CHOL in raft-like membranes leads to strongly packed and rigid bilayers. We also find that the simulated raft bilayers are characterized by nanoscale lateral heterogeneity, though the slow lateral diffusion renders the interpretation of the observed lateral heterogeneity more difficult. The findings reveal aspects of the role of favored (specific) lipid–lipid interactions within rafts and clarify the prominent role of CHOL in altering the properties of the membrane locally in its neighborhood. Also, we show that the presence of PSM and CHOL in rafts leads to intriguing lateral pressure profiles that are distinctly different from corresponding profiles in nonraft-like membranes. The results propose that the functioning of certain classes of membrane proteins is regulated by changes in the lateral pressure profile, which can be altered by a change in lipid content.

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Lipid interactions with bacterial channels: fluorescence studies

Cited by 14 publications

References 18 publications

Properties of the Mechanosensitive Channel MscS Pore Revealed by Tryptophan Scanning Mutagenesis

Properties of the Mechanosensitive Channel MscS Pore Revealed by Tryptophan Scanning Mutagenesis

Regulation of voltage-gated potassium channels by PI(4,5)P2

Assessing the Nature of Lipid Raft Membranes

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