Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.16.2 (PCP)
 3,761 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this study an enzyme-linked immunosorbent assay (ELISA) was developed to detect the stress protein Hsp70 in the green alga Raphidocelis subcapitata. Using this ELISA, the response to a variety of pollutants, including ZnCl2, SeO2 (heavy metals), lindane (organochlorine pesticide), pentachlorophenol (PCP, chlorinated hydrocarbon insecticide and fungicide), carbaryl (carbamate pesticide) and sodium dodecyl sulphate (SDS; surfactant) was tested. Our results show that Hsp 70 is produced in a dose-dependent way in response to most chemicals investigated (except PCP) and at concentrations below the range of classical cytotoxicity testing (i.e. growth inhibition, lethality). Still, the potential to induce Hsp70 varied among the pollutants tested, the heavy metals ZnCl2 and SeO2 being the strongest inducers of Hsp70. Combined with the existing literature, these results indicate that Hsp70 in R. subcapitata is a sensitive biomarker for a wide range of environmental pollutants.
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PMID:Dose-dependent induction of heat shock protein 70 synthesis in Raphidocelis subcapitata following exposure to different classes of environmental pollutants. 1509 1

A membrane-associated 3,5-dichlorophenol reductive dehalogenase was isolated from Desulfitobacterium frappieri PCP-1. The highest dehalogenase activity was observed with the biomass cultured at 22 degrees C, compared to 30 and 37 degrees C, where the cell suspensions were 2.2 and 9.6 times less active, respectively. The reductive dehalogenase was purified 12.7-fold to apparent homogeneity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single band with an apparent molecular mass of 57 kDa. Its dechlorinating activity was not inhibited by sulfate and nitrate but was completely inhibited by 2.5 mM sulfite and 10 mM KCN. A mixture of iodopropane and titanium citrate caused a light-reversible inhibition of the dechlorinating activities, suggesting the involvement of a corrinoid cofactor. Several polychlorophenols were dechlorinated at the meta and para positions. The apparent K(m) for 3,5-dicholorophenol was 49.3 +/- 3.1 microM at a methyl viologen concentration of 2 mM. Six internal tryptic peptides were sequenced by mass spectrometry. One open reading frame (ORF) was found in the Desulfitobacterium hafniense genome containing these peptide sequences. This ORF corresponds to a gene coding for a CprA-type reductive dehalogenase. The corresponding ORF (named cprA5) in D. frappieri PCP-1 was cloned and sequenced. The cprA5 gene codes for a 548-amino-acid protein that contains a twin-arginine-type signal for secretion. The gene product has a cobalamin binding site motif and two iron-sulfur binding motifs and shows 66% identity (76 to 77% similarity) with some tetrachloroethene reductive dehalogenases. This is the first CprA-type reductive dehalogenase that can dechlorinate chlorophenols at the meta and para positions.
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PMID:Purification, cloning, and sequencing of a 3,5-dichlorophenol reductive dehalogenase from Desulfitobacterium frappieri PCP-1. 1529 82

FXYD domain-containing proteins are tissue-specific regulators of the Na,K-ATPase that have been shown to have significant physiological implications. Information about the sites of interaction between some FXYD proteins and subunits of the Na,K-ATPase is beginning to emerge. We previously identified an FXYD protein in plasma membranes from shark rectal gland cells and demonstrated that this protein (FXYD10) modulates shark Na,K-ATPase activity. The present study was undertaken to identify the location of the C-terminal domain of FXYD10 on the alpha-subunit of Na,K-ATPase, using covalent cross-linking combined with proteolytic cleavage. Treatment of Na,K-ATPase-enriched membranes with the homobifunctional thiol cross-linker 1,4-bismaleimidyl-2,3-dihydroxybutane resulted in cross-linking of FXYD10 to the alpha-subunit. Cross-linking was not affected by preincubation with sodium or potassium but was significantly reduced after pre-incubation with the non-hydrolyzable ATP analog beta,gamma-methyleneadenosine 5'-triphosphate (AMP-PCP). A peptic assay was developed, in which pepsin treatment of Na,K-ATPase at low pH resulted in extensive cleavage of the alpha-subunit while FXYD10 was left intact. Proteolytic fragments of control and cross-linked preparations were isolated by immunoprecipitation and analyzed by gel electrophoresis. A proteolytic fragment containing FXYD10 cross-linked to a fragment from the alpha-subunit could be localized on SDS gels. Sequencing of this fragment showed the presence of FXYD10 as well as a fragment within the A domain of the alpha-subunit comprising 33 amino acids, including a single Cys residue, Cys254. Thus, regulation of Na,K-ATPase by FXYD10 occurs in part via cytoplasmic interaction of FXYD10 with the A domain of the shark alpha-subunit.
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PMID:Interaction of FXYD10 (PLMS) with Na,K-ATPase from shark rectal glands. Close proximity of Cys74 of FXYD10 to Cys254 in the a domain of the alpha-subunit revealed by intermolecular thiol cross-linking. 1591 65

Radioligand binding, photoaffinity labeling, and docking and molecular dynamics were used to characterize the tricyclic antidepressant (TCA) binding sites in the nicotinic acetylcholine receptor (nAChR). Competition experiments indicate that the noncompetitive antagonist phencyclidine (PCP) inhibits [3H]imipramine binding to resting (closed) and desensitized nAChRs. [3H]2-azidoimipramine photoincorporates into each subunit from the desensitized nAChR with approximately 25% of the labeling specifically inhibited by TCP (a PCP analog), whereas no TCP-inhibitable labeling was observed in the resting (closed) state. For the desensitized nAChR and within the alpha subunit, the majority of specific [3H]2-azidoimipramine labeling mapped to a approximately 20 kDa Staphylococcus aureus V8 protease fragment (alphaV8-20; Ser173-Glu338). To further map the labeling site, the alphaV8-20 fragment was further digested with endoproteinase Lys-C and resolved by Tricine SDS-PAGE. The principal labeled fragment (11 kDa) was further purified by rpHPLC and subjected to N-terminal sequencing. Based on the amino terminus (alphaMet243) and apparent molecular weight, the 11 kDa fragment contains the channel lining M2 segment. Finally, docking and molecular dynamics results indicate that imipramine and PCP interact preferably with the M2 transmembrane segments in the middle of the ion channel. Collectively, these results are consistent with a model where PCP and TCA bind to overlapping sites within the lumen of the Torpedo nAChR ion channel.
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PMID:Identifying the binding site(s) for antidepressants on the Torpedo nicotinic acetylcholine receptor: [3H]2-azidoimipramine photolabeling and molecular dynamics studies. 1881 47


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