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Plasma Membrane Fraction (plasma + membrane_fraction)
Selected AbstractsThe proteome survey of an electricity-generating organ (Torpedo californica electric organ)PROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 4 2007Javad Nazarian Abstract Torpedo californica is a species in class Chondrichthyes. Electric rays have evolved the electric organ, which is similar to the mammalian neuromuscular junction (NMJ). Here, we took a combined cDNA sequencing and proteomic approach to define the molecular constituents of the T. californica electric organ. For soluble proteins, 2-DE was used and 224 protein spots were mapped. Plasma membrane fractions were analyzed using the shotgun approach (LC-MS/MS). A Torpedo cDNA library was constructed and 607 cDNA clones were sequenced. Identification of electric organ proteins was done using cross-species comparisons, and a custom database was constructed from cDNA translations. We unambiguously identified 121 proteins and transcripts, 103 of which were novel additions to the existing databases of Torpedo fish. Fifteen proteins of known function, but not previously associated with either the electroplaque or NMJ, were present at high abundance. These included the heat shock and oxidative stress proteins, annexin V (calelectrin), and plectin 1. Most interesting were the unambiguous matches to 11 human ORFs of unknown function, including four potential RNA splicing proteins, a vacuolar sorting protein, and a tetraspanin containing protein. This analysis identified proteins that may play a role in the higher vertebrate neuromuscular junction or other electrical synapses. [source] Temperature-dependent localization of GPI-anchored intestinal alkaline phosphatase in model rafts,JOURNAL OF MOLECULAR RECOGNITION, Issue 6 2007Marie-Cécile Giocondi Abstract In plasma membranes, most of glycosylphosphatidylinositol (GPI)-anchored proteins would be associated with rafts, a category of ordered microdomains enriched in sphingolipids and cholesterol (Ch). They would be also concentrated in the detergent resistant membranes (DRMs), a plasma membrane fraction extracted at low temperature. Preferential localization of GPI-anchored proteins in these membrane domains is essentially governed by their high lipid order, as compared to their environment. Changes in the temperature are expected to modify the membrane lipid order, suggesting that they could affect the distribution of GPI-anchored proteins between membrane domains. Validity of this hypothesis was examined by investigating the temperature-dependent localization of the GPI-anchored bovine intestinal alkaline phophatase (BIAP) into model raft made of palmitoyloleoylphosphatidylcholine/sphingomyelin/cholesterol (POPC/SM/Chl) supported membranes. Atomic force microscopy (AFM) shows that the inserted BIAP is localized in the SM/Chl enriched ordered domains at low temperature. Above 30°C, BIAP redistributes and is present in both the ,fluid' POPC enriched and the ordered SM/Chl domains. These data strongly suggest that in cells the composition of plasma membrane domains at low temperature differs from that at physiological temperature. Copyright © 2007 John Wiley & Sons, Ltd. [source] High salt-treatment-induced Na+ extrusion and low salt-treatment-induced Na+ accumulation in suspension-cultured cells of the mangrove plant, Bruguiera sexangulaPLANT CELL & ENVIRONMENT, Issue 10 2001M. Kura-Hotta Abstract A suspension-cultured cell strain of the mangrove plant (Bruguiera sexangula) was established from a callus culture and maintained in an amino acid medium in the absence of NaCl. NaCl non-adapted cells were transferred to media containing 0,200 mm NaCl. The initial growth rate decreased gradually with increasing salt concentrations. However, at up to 150 mm NaCl, cell number growth at the highest point was almost the same as that at lower salt concentrations. Cells even continued to grow in the presence of 200 mm NaCl. Cells incubated in a medium containing 50 mm NaCl for 3 weeks accumulated Na+, while those incubated in 150 mm NaCl for 2 d showed only a transient increase in Na+ and Cl, concentrations. In the latter treatment, the intracellular concentration of Na+ returned to the original low level within 2 weeks. It took a longer time for Cl, to return to its original level. As a result, the Na+ and Cl, concentrations in cells cultured with 50 mm NaCl were much larger than those in cells cultured with 150 mm NaCl. The intracellular distribution of ions after transfer to the medium containing 150 mm NaCl was analysed by isolating the vacuoles. Treatment with amiloride, an inhibitor of the Na+/H+ antiporter, suppressed the recovery of Na+ to the original level in the cells. Treatment with 150 mm NaCl for 3 d stimulated the activities of both the vanadate-dependent H+ -ATPase and the Na+/H+ antiporter in the plasma membrane fraction. [source] Characterization of human sperm N -acetylglucosaminidaseINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 3 2008S. L. Perez Martinez Summary N -acetylglucosaminidase (NAG) is particularly active in mammalian spermatozoa and appears to be involved in fertilization. Although it is assumed that this enzyme is acrosomal, previous results from our laboratory suggest the presence of NAG at the sperm plasma membrane level. The present study attempted to analyse the subcellular distribution of this enzyme in human spermatozoa. Sperm were incubated under different conditions and NAG activity measured in the soluble extracts and cell pellets using a specific fluorometric substrate. A significant proportion of NAG activity was released when sperm were incubated in culture medium, suggesting a weak association with the plasma membrane. This location was confirmed by western blot analysis of plasma membrane fractions and immunofluorescence on non-permeabilized sperm, which showed a positive signal mainly on the acrosomal domain. The distribution of NAG activity between plasma membrane and acrosome was analysed after cell disruption by freezing and thawing. Triton X-100 stimulated sperm and epididymal NAG activity but not the enzyme obtained from other sources. In addition, biotinylated human recombinant NAG was able to bind to human sperm. Finally, after sperm incubation under capacitating conditions, NAG total activity increased and the sperm enzyme lost its ability to be stimulated by Triton X-100. The possible connection of these results with sperm maturation, capacitation and NAG participation in primary binding to the zona pellucida, was discussed. [source] | ||||||||||