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J Phys Chem B . 2012 Feb 15; Horst R, Stanczak P, Serrano P, Wüthrich K ABSTRACT: Aqueous solutions of the detergent Fos-10 (n-decylphosphocholine) without and with addition of the integral membrane protein (IMP) OmpX (outer membrane protein X) have been characterized using pulsed field gradient-stimulated echo (PFG-STE) NMR experiments for measurements of translational diffusion coefficients. Effective diffusion coefficients for Fos-10 micelles in the absence of OmpX were obtained by observation of NMR signals from 10-bromodecan-1-ol that had been inserted into the micelles, and in the presence of OmpX by NMR observation of the protein. Read more from the original source: Applications are invited for the post of Junior Research Fellow (JRF) under a DST Project entitled “Structural Characterization of Integral Membrane Proteins with Proteolytic activity from Pathogenic Bacterial species” funded … Read more here: J. Am. Chem. Soc. In Journal of the American Chemical Society, Vol. 131, No. 35. (12 August 2009), pp. 12650-12656. We measured the lateral mobility of integral membrane proteins reconstituted in giant unilamellar vesicles (GUVs), using fluorescence correlation spectroscopy. Receptor, channel, and transporter proteins with 1?36 transmembrane segments (lateral radii ranging from 0.5 to 4 nm) and a α-helical peptide (radius of 0.5 nm) were fluorescently labeled and incorporated into GUVs. At low protein-to-lipid ratios (i.e., 10?100 proteins per ?m2 of membrane surface), the diffusion coefficient D displayed a weak dependence on the hydrodynamic radius (R) of the proteins [D scaled with ln(1/R)], consistent with the Saffman-Delbru?ck model. At higher protein-to lipid ratios (up to 3000 ?m?2), the lateral diffusion coefficient of the molecules decreased linearly with increasing the protein concentration in the membrane. The implications of our findings for protein mobility in biological membranes (protein crowding of ?25,000 ?m?2) and use of diffusion measurements for protein geometry (size, oligomerization) determinations are discussed. We measured the lateral mobility of integral membrane proteins reconstituted in giant unilamellar vesicles (GUVs), using fluorescence correlation spectroscopy. Receptor, channel, and transporter proteins with 1?36 transmembrane segments (lateral radii ranging from 0.5 to 4 nm) and a α-helical peptide (radius of 0.5 nm) were fluorescently labeled and incorporated into GUVs. At low protein-to-lipid ratios (i.e., 10?100 proteins per ?m2 of membrane surface), the diffusion coefficient D displayed a weak dependence on the hydrodynamic radius (R) of the proteins [D scaled with ln(1/R)], consistent with the Saffman-Delbru?ck model. At higher protein-to lipid ratios (up to 3000 ?m?2), the lateral diffusion coefficient of the molecules decreased linearly with increasing the protein concentration in the membrane. The implications of our findings for protein mobility in biological membranes (protein crowding of ?25,000 ?m?2) and use of diffusion measurements for protein geometry (size, oligomerization) determinations are discussed. J. Am. Chem. Soc. In Journal of the American Chemical Society, Vol. 131, No. 35. (12 August 2009), pp. 12650-12656. We measured the lateral mobility of integral membrane proteins reconstituted in giant unilamellar vesicles (GUVs), using fluorescence correlation spectroscopy. Receptor, channel, and transporter proteins with 1?36 transmembrane segments (lateral radii ranging from 0.5 to 4 nm) and a α-helical peptide (radius of 0.5 nm) were fluorescently labeled and incorporated into GUVs. At low protein-to-lipid ratios (i.e., 10?100 proteins per ?m2 of membrane surface), the diffusion coefficient D displayed a weak dependence on the hydrodynamic radius (R) of the proteins [D scaled with ln(1/R)], consistent with the Saffman-Delbru?ck model. At higher protein-to lipid ratios (up to 3000 ?m?2), the lateral diffusion coefficient of the molecules decreased linearly with increasing the protein concentration in the membrane. The implications of our findings for protein mobility in biological membranes (protein crowding of ?25,000 ?m?2) and use of diffusion measurements for protein geometry (size, oligomerization) determinations are discussed. We measured the lateral mobility of integral membrane proteins reconstituted in giant unilamellar vesicles (GUVs), using fluorescence correlation spectroscopy. Receptor, channel, and transporter proteins with 1?36 transmembrane segments (lateral radii ranging from 0.5 to 4 nm) and a α-helical peptide (radius of 0.5 nm) were fluorescently labeled and incorporated into GUVs. At low protein-to-lipid ratios (i.e., 10?100 proteins per ?m2 of membrane surface), the diffusion coefficient D displayed a weak dependence on the hydrodynamic radius (R) of the proteins [D scaled with ln(1/R)], consistent with the Saffman-Delbru?ck model. At higher protein-to lipid ratios (up to 3000 ?m?2), the lateral diffusion coefficient of the molecules decreased linearly with increasing the protein concentration in the membrane. The implications of our findings for protein mobility in biological membranes (protein crowding of ?25,000 ?m?2) and use of diffusion measurements for protein geometry (size, oligomerization) determinations are discussed.
Int J Cell Biol. 2012; 2012: 121424 The establishment and maintenance of the polarized epithelial phenotype require a characteristic organization of the cytoskeletal components. There are many cellular effectors involved in the regulation of the cytoskeleton of epithelial cells. Recently, modifications in the plasma membrane potential (PMP) have been suggested to participate in the modulation of the cytoskeletal organization of epithelia. Here, we review evidence showing that changes in the PMP of diverse epithelial cells promote characteristic modifications in the cytoskeletal organization, with a focus on the actin cytoskeleton. The molecular paths mediating these effects may include voltage-sensitive integral membrane proteins and/or peripheral proteins sensitive to surface potentials. The voltage dependence of the cytoskeletal organization seems to have implications in several physiological processes, including epithelial wound healing and apoptosis. Visit link:
Int J Cell Biol. 2012; 2012: 121424 The establishment and maintenance of the polarized epithelial phenotype require a characteristic organization of the cytoskeletal components. There are many cellular effectors involved in the regulation of the cytoskeleton of epithelial cells. Recently, modifications in the plasma membrane potential (PMP) have been suggested to participate in the modulation of the cytoskeletal organization of epithelia. Here, we review evidence showing that changes in the PMP of diverse epithelial cells promote characteristic modifications in the cytoskeletal organization, with a focus on the actin cytoskeleton. The molecular paths mediating these effects may include voltage-sensitive integral membrane proteins and/or peripheral proteins sensitive to surface potentials. The voltage dependence of the cytoskeletal organization seems to have implications in several physiological processes, including epithelial wound healing and apoptosis. Read more from the original source:
Biopolymers. 2012 Feb 3; Asparagine-linked glycosylation is an essential and highly conserved protein modification reaction. In eukaryotes, oligosaccharyl transferase (OT), a multisubunit membrane-associated enzyme complex, catalyzes this reaction in newly synthesized proteins. In Saccharomyces cerevisiae, OT consists of nine non-identical membrane proteins. Ost4p, the smallest subunit, bridges the catalytic subunit Stt3p with Ost3p. Mutation of transmembrane residues 18-24 in Ost4p has negative effect on OT activity, disrupts the Stt3p-Ost4p-Ost3p complex, results in temperature-sensitive phenotype, and hypoglycosylation. Heterologous expression and purification of integral membrane proteins (IMPs) are the bottleneck in membrane protein research. We report the cloning, successful overexpression and purification of recombinant Ost4p with a novel but simple method producing milligram quantities of pure protein. GB1 protein was found to be the most suitable tag for the large scale production of Ost4p. The cleavage of Ost4p conveniently leaves GB1 protein in solution eliminating further purification. The precipitated pure Ost4p is reconstituted in appropriate membrane mimetic. The recombinant protein is highly helical as indicated by the far-UV CD spectrum. The well-dispersed HSQC spectrum indicates that this minimembrane protein is well folded. The successful production of pure recombinant Ost4p with a novel yet simple method may have important ramification for the production of other membrane proteins. © 2012 Wiley Periodicals, Inc. Biopolymers, 2012. The rest is here: (NYCOMPS) an NIH-supported PSI: Biology Center for Membrane Protein Structure Determination. Protein production … postdoctoral experience. Expertise with integral membrane proteins and/or x-ray crystallography would be helpful, but… Read more here: Abstract Micro integral membrane protein (MIMP) has been shown to adhere to mucin and antagonize the adhesion of enteropathogenic Escherichia coli (EPEC) to epithelial cells, however, the mechanism has not been fully … Follow this link: highly purify membrane proteins (e.g. integral membrane proteins, GPCRs, ion channels and membrane-associated … hands-on small-scale and large-scale purification of membrane proteins using state-of-the-art methods and… Read more here: highly purify membrane proteins (e.g. integral membrane proteins, GPCRs, ion channels and membrane-associated … hands-on small-scale and large-scale purification of membrane proteins using state-of-the-art methods and… Read more here: |
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