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11.
Patrizzia Mastromatteo-Alberga Fabiola Placeres-Uray Marcelo A. Alfonzo-González Ramona Gonzalez de Alfonzo Itala Lippo de Becemberg 《Journal of receptor and signal transduction research》2016,36(3):278-287
Muscarinic antagonists, via muscarinic receptors increase the cAMP/cGMP levels at bovine tracheal smooth muscle (BTSM) through the inhibition of phosphodiesterases (PDEs), displaying a similar behavior of vinpocetine (a specific-PDE1 inhibitor). The presence of PDE1 hydrolyzing both cyclic nucleotides in BTSM strips was revealed. Moreover, a vinpocetine and muscarinic antagonists inhibited PDE1 located at plasma membranes (PM) fractions from BTSM showing such inhibition, an M2AChR pharmacological profile. Therefore, a novel Ca2+/CaM dependent and vinpocetine inhibited PDE1 was purified and characterized at PM fractions from BTSM. This PDE1 activity was removed from PM fractions using a hypotonic buffer and purified some 38 fold using two columns (Q-Sepharose and CaM-agarose). This PDE1 was stimulated by CaM and inhibited by vinpocetine showing two bands in PAGE-SDS (56, 58?kDa) being the 58?kDa identified as PDE1A by Western blotts. This PDE1A activity was assayed with [3H]cGMP and [3H]cAMP exhibiting a higher affinity as Km (μM) for cGMP than cAMP but being close values with Vmax cAMP/cGMP ratio of 1.5. The co-factor Mg2+ showed similar K(A) (mM) for both cyclic nucleotides. Vinpocetine showed similar inhibition concentration 50% (IC50 of 4.9 and 4.6?μM) for cAMP and cGMP, respectively. CaM stimulated the cyclic nucleotides hydrolysis by PDE1A exhibiting similar activation constant as K(CaM), in nM range. The original finding was the identification and purification of a vinpocetine and muscarinic antagonist-inhibited and CaM-activated PM-bound PDE1A, linked to M2AChR. A model of this novel signal transducing cascade for the regulation of cyclic nucleotides levels at BTSM is proposed. 相似文献
12.
The GDH (NADPH) mutant strain of has sizable pools of glutamine and glutamate under ammonium-limited conditions for which requires an elevated glutamine synthetase activity. Glutamine in the pre ence of 2-oxoglutarate, stimulated nicotinamide nucleotide oxidation by crude and purified extracts of the strain and led to a reductant dependent formation of two molecules of glutamate. Aminooxyacetate did not have any effect on the reaction, whereas azaserine inhibited it completely. It is concluded that in glutamine synthetase and glutamate synthase are responsible for the assimilation of low ammonium concentrations. 相似文献
13.
Vicente Bermúdez Silvia S. Antollini Gaspar A. Fernández Nievas Marta I. Avelda?o Francisco J. Barrantes 《Journal of lipid research》2010,51(9):2629-2641
The nicotinic acetylcholine receptor (AChR) is in intimate contact with the
lipids in its native membrane. Here we analyze the possibility that it is the
intrinsic properties of the AChR that determine its partition into a given lipid
domain. Torpedo AChR or a synthetic peptide corresponding to
the AChR γM4 segment (the one in closer contact with lipids) was
reconstituted into “raft”-containing model
membranes. The distribution of the AChR was assessed by Triton X-100 extraction
in combination with fluorescence studies, and lipid analyses were performed on
each sample. The influence of rapsyn, a peripheral protein involved in AChR
aggregation, was studied. Raft-like domain aggregation was also
studied using membranes containing the ganglioside GM1 followed by GM1
crosslinking. The γM4 peptide displays a marked preference for
raft-like domains. In contrast, AChR alone or in the
presence of rapsyn or ganglioside aggregation exhibits no such preference for
raft-like domains, but it does cause a significant reduction in the total amount
of these domains. The results indicate that the distribution of the AChR in
lipid domains cannot be due exclusively to the intrinsic physicochemical
properties of the protein and that there must be an external signal in native
cell membranes that directs the AChR to a specific membrane domain. 相似文献
14.
E M Kosower 《FEBS letters》1983,157(1):144-146
A molecular model for the bilayer helices of the acetylcholine receptor is constructed from the 7 channel elements and the 17 hydrophobic helices of the 5 protein subunits. The acetylcholine binding site and the opening to the ion channel are included. 相似文献
15.
M R Hanley 《Biochemical and biophysical research communications》1978,82(1):392-401
The phospholipolytic neurotoxin from , crotoxin, is able to produce a dose- and time-dependent block of carbachol-stimulated 22Na efflux from pre-loaded excitable vesicles. The blocking activity is dependent on calcium and is abolished by chemical modification with p-bromophenacyl bromide. The isolated basic subunit, crotoxin B, produces an identical block, whereas the isolated acidic subunit, crotoxin A, has no detectable effect. Neither crotoxin nor crotoxin B antagonizes the binding of to purified acetylcholine receptor, although, at high concentrations, they antagonize its binding to acetylcholine receptor-rich membrane fragments. Certain phospholipase A2 enzymes and the fatty acid products of their digestion can mimic the crotoxin action. It is therefore suggested that, although considered a pre-synaptic neurotoxin, crotoxin can have post-synaptic effects, possibly mediated by its endogeneous phospholipase A2 activity. 相似文献
16.
Endocytosed nicotinic acetylcholine receptors (CHRN) are degraded via macroautophagy/autophagy during atrophic conditions and are accompanied by the autophagic regulator protein SH3GLB1. The present study addressed the functional role of SH3GLB1 on CHRN trafficking and its implementation. We found an augmented ratio of total SH3GLB1 to threonine-145 phosphorylated SH3GLB1 (SH3GLB1:p-SH3GLB1) under conditions of increased CHRN vesicle numbers. Overexpression of T145 phosphomimetic (T145E) and phosphodeficient (T145A) mutants of SH3GLB1, was found to either slow down or augment the processing of endocytic CHRN vesicles, respectively. Co-expression of the early endosomal orchestrator RAB5 largely rescued the slow processing of endocytic CHRN vesicles induced by T145E. SH3GLB1 phosphomutants did not modulate the expression or colocalization of RAB5 with CHRN vesicles, but instead altered the expression of RAB5 activity regulators. In summary, these findings suggest that SH3GLB1 controls CHRN endocytic trafficking in a phosphorylation- and RAB5-dependent manner at steps upstream of autophagosome formation. 相似文献
17.
18.
The A12 acetylcholinesterase and polypeptide composition of electric organ basal lamina of Electrophorus and some Torpedinae fishes 总被引:1,自引:0,他引:1
Basal lamina (BL) of Torpedo, Discopyge and Electrophorus electric organs was purified in order to establish polypeptide composition and association with acetylcholinesterase (AChE). Results indicate that BL presents a distinct peptide pattern and that the A12 form of AChE is directly attached to it. Comparison of the species studied demonstrated similarities both in polypeptide composition and AChE content of the purified BL. Extractions of BL with solutions of high ionic strength, guanidine-HCl and acetic acid indicated the differential solubilization of various domains of BL polypeptides. 相似文献
19.
The role of adenosine 5'-triphosphate (ATP) and P2Y(1) nucleotide receptor in potentiating agrin-induced acetylcholine receptor (AChR) aggregation is being demonstrated in a co-culture system of NG108-15 cell, a mouse neuroblastoma X rat glioma hybrid cell line that resembles spinal motor neuron, with C2C12 myotube. In the co-cultures, antagonized P2Y(1) receptors showed a reduction in NG108-15 cell-induced AChR aggregation. Parallel to this observation, cultured NG108-15 cell secreted ATP into the conditioned medium in a time-dependent manner. Enhancement of ATP release from the cultured NG108-15 cells by overexpression of active mutants of small GTPases increased the aggregation of AChRs in co-culturing with C2C12 myotubes. In addition, ecto-nucleotidase was revealed in the co-culture, which rapidly degraded the applied ATP. These results support the notion that ATP has a role in directing the formation of post-synaptic apparatus in vertebrate neuromuscular junctions. 相似文献
20.
Richard Grantham 《FEBS letters》1980,121(2):193-199