A cytotoxic sulfhydryl reagent that inhibits several subcellular metabolic systems and is used as a tool in cellular physiology.

Transport of fluid by lens epithelium. (1/190)

We report for the first time that cultured lens epithelial cell layers and rabbit lenses in vitro transport fluid. Layers of the alphaTN4 mouse cell line and bovine cell cultures were grown to confluence on permeable membrane inserts. Fluid movement across cultured layers and excised rabbit lenses was determined by volume clamp (37 degrees C). Cultured layers transported fluid from their basal to their apical sides against a pressure head of 3 cmH2O. Rates were (in microliter. h-1. cm-2) 3.3 +/- 0.3 for alphaTN4 cells (n = 27) and 4.7 +/- 1.0 for bovine layers (n = 6). Quinidine, a blocker of K+ channels, and p-chloromercuribenzenesulfonate and HgCl2, inhibitors of aquaporins, inhibited fluid transport. Rabbit lenses transported fluid from their anterior to their posterior sides against a 2.5-cmH2O pressure head at 10.3 +/- 0.62 microliter. h-1. lens-1 (n = 5) and along the same pressure head at 12.5 +/- 1.1 microliter. h-1. lens-1 (n = 6). We calculate that this flow could wash the lens extracellular space by convection about once every 2 h and therefore might contribute to lens homeostasis and transparency.  (+info)

Transmembrane segment 5 of the Glut1 glucose transporter is an amphipathic helix that forms part of the sugar permeation pathway. (2/190)

Transmembrane segment 5 of the Glut1 glucose transporter has been proposed to form an amphipathic transmembrane helix that lines the substrate translocation pathway (Mueckler, M., Caruso, C., Baldwin, S. A., Panico, M., Blench, I., Morris, H. R., Allard, W. J., Lienhard, G. E., and Lodish, H. F. (1985) Science 229, 941-945). This hypothesis was tested using cysteine-scanning mutagenesis in conjunction with the membrane-impermeant, sulfhydryl-specific reagent, p-chloromercuribenzenesulfonate (pCMBS). A series of 21 mutants was created from a fully functional, cysteine-less, parental Glut1 molecule by changing each residue within putative transmembrane segment 5 to cysteine. Each mutant was then expressed in Xenopus oocytes and its steady-state protein level, 2-deoxyglucose uptake activity, and sensitivity to pCMBS were measured. All 21 mutants exhibited measurable transport activity, although several of the mutants exhibited reduced activity due to a corresponding reduction in steady-state protein. Six of the amino acid side chains within transmembrane segment 5 were clearly accessible to pCMBS in the external medium, as determined by inhibition of transport activity, and a 7th residue showed inhibition that lacked statistical significance because of the extremely low transport activity of the corresponding mutant. All 7 of these residues were clustered along one face of a putative alpha-helix, proximal to the exoplasmic surface of the plasma membrane. These results comprise the first experimental evidence for the existence of an amphipathic transmembrane alpha-helix in a glucose transporter molecule and strongly suggest that transmembrane segment 5 of Glut1 forms part of the sugar permeation pathway.  (+info)

Identification of the amine-polyamine-choline transporter superfamily 'consensus amphipathic region' as the target for inactivation of the Escherichia coli GABA transporter GabP by thiol modification reagents. Role of Cys-300 in restoring thiol sensitivity to Gabp lacking Cys. (3/190)

The Escherichia coli gamma-aminobutyric acid transporter GabP (gab permease) contains a functionally significant cysteine residue (Cys-300) within its consensus amphipathic region (CAR), a putative channel-forming structure that extends out of transmembrane helix 8 and into the adjoining cytoplasmic loop 8-9 of transporters from the amine-polyamine-choline (APC) superfamily. Here we show that of the five cysteine residues (positions 158, 251, 291, 300 and 443) in the E. coli GabP, Cys-300 is the one that renders the transport activity sensitive to inhibition by thiol modification reagents: whereas substituting Ala for Cys-300 mimics the inhibitory effect of thiol modification, substituting Ala at position 158, 251, 291 or 443 preserves robust transport activity and confers no resistance to thiol inactivation; and whereas the robustly active Cys-300 single-Cys mutant is fully sensitive to thiol modification, other single-Cys mutants (Cys at 158, 251, 291 or 443) exhibit kinetically compromised transport activities that resist further chemical inactivation by thiol reagents. The present study reveals additionally that Cys-300 exhibits (1) sensitivity to hydrophobic thiol reagents, (2) general resistance to bulky (fluorescein 5-maleimide) and/or charged {2-sulphonatoethyl methanethiosulphonate or [2-(trimethylammonium)ethyl] methanethiosulphonate} thiol reagents and (3) a peculiar sensitivity to p-chloromercuribenzenesulphonate (PCMBS). The accessibility of PCMBS to Cys-300 (located midway through the lipid bilayer) might be related to the structural similarity that it shares with guvacine (1, 2,3,6-tetrahydro-3-pyridinecarboxylic acid), a transported GabP substrate. These structural requirements for thiol sensitivity provide the first chemical evidence consistent with channel-like access to the polar surface of the CAR, a physical configuration that might provide a basis for understanding how this region impacts the function of APC transporters generally [Closs, Lyons, Kelly and Cunningham (1993) J. Biol. Chem. 268, 20796-20800] and the gab permease particularly [Hu and King (1998) Biochem. J. 300, 771-776].  (+info)

Melibiose carrier of Escherichia coli: use of cysteine mutagenesis to identify the amino acids on the hydrophilic face of transmembrane helix 2. (4/190)

The melibiose carrier from Escherichia coli is a galactoside-cation symporter. Based on both experimental evidence and hydropathy analysis, 12 transmembrane helices have been assigned to this integral membrane protein. Transmembrane helix 2 contains several charged and polar amino acids that have been shown to be essential for the cation-coupled transport of melibiose. Starting with the cysteine-less melibiose carrier, we have individually substituted cysteine for amino acids 39-66, which includes the proposed transmembrane helix 2. In the resulting derivative carriers, we measured the transport of melibiose, determined the effect of the hydrophilic sulfhydryl reagent, p-chloromercuribenzenesulfonic acid (PCMBS), on transport in intact cells and inside out vesicles, and examined the ability of melibiose to protect the carrier from inactivation by the sulfhydryl reagent. We found a set of seven positions in which the reaction with the sulfhydryl reagent caused partial or complete loss of carrier function measured in intact cells or inside-out vesicles. The presence of melibiose protected five of these positions from reaction with PCMBS. The reaction of two additional positions with PCMBS resulted in the partial loss of transport function only in inside-out vesicles. Melibiose protected these two positions from reaction with the reagent. Together, the PCMBS-sensitive sites and charged residues assigned to helix 2 form a cluster of amino acids that map in three rows with each row comprised of every fourth residue. This is the pattern expected of residues that are part of an alpha-helical structure and thus the rows are tilted at an angle of 25 degrees to the helical axis. We suggest that these residues line the path of melibiose and its associated cation through the carrier.  (+info)

Roles of Gln81 and Cys80 in catalysis by glycosylphosphatidylinositol-phospholipase C from Trypanosoma brucei. (5/190)

Glycosylphosphatidylinositol-specific phospholipase C (GPtdIns-PLC) is found in the protozoan parasite Trypanosoma brucei. A region of protein sequence similarity exists between the protozoan enzyme and eubacterial phosphatidylinositol-phospholipases C. The functional relevance of Cys80 and Gln81 of GPtdIns-PLC, both in this region, was tested with a panel of mutations at each position. Gln81Glu, Gln81Ala, Gln81Gly, Gln81Lys and Gln81Leu mutants were inactive. Cleavage of GPtdIns was detectable in Gln81Asn, although the specific activity decreased 500-fold, and kcat was reduced 50-fold. Thus an amide side-chain at residue 81 is essential for catalysis by GPtdIns-PLC. Sulfhydryl reagents inactivate GPtdIns-PLC, suggesting that a Cys could be close to the enzyme active site. Surprisingly, p-chloromercuriphenyl sulfonate (p-CMPS) is significantly more potent than N-ethylmaleimide, the less bulky compound. This knowledge prompted us to test whether replacement of Cys80 with an amino acid possessing a bulky side-chain would inactivate GPtdIns-PLC: Cys80Ala, Cys80Thr, Cys80Phe, Cys184Ala, and Cys269-270-273Ser were constructed for that purpose. Cys80Phe lacked enzyme activity, while Cys80Ala, Cys80Thr and Cys269-270-273Ser retained 33 to 100% of wild-type activity. Interestingly, the Cys80Ala and Cys80Thr mutants became resistant to p-CMPS, as predicted if the sulfhydryl reagent reacted with Cys80 in the wild-type enzyme to form a cysteinyl mercurylphenylsulfonate moiety, a bulky adduct that inactivated GPtdIns-PLC, similar to the Cys80Phe mutation. We conclude that a bulky side-chain (or adduct) at position 80 of GPtdIns-PLC abolishes enzyme activity. Together, these observations place Cys80 and Gln81 at, or close to, the active site of GPtdIns-PLC from T. brucei.  (+info)

Gamma-aminobutyric acid increases the water accessibility of M3 membrane-spanning segment residues in gamma-aminobutyric acid type A receptors. (6/190)

Gamma-aminobutyric acid type A (GABA(A)) receptors are members of the ligand-gated ion channel gene superfamily. Using the substituted cysteine accessibility method, we investigated whether residues in the alpha(1)M3 membrane-spanning segment are water-accessible. Cysteine was substituted, one at a time, for each M3 residue from alpha(1)Ala(291) to alpha(1)Val(307). The ability of these mutants to react with the water-soluble, sulfhydryl-specific reagent pCMBS(-) was assayed electrophysiologically. Cysteines substituted for alpha(1)Ala(291) and alpha(1)Tyr(294) reacted with pCMBS(-) applied both in the presence and in the absence of GABA. Cysteines substituted for alpha(1)Phe(298), alpha(1)Ala(300), alpha(1)Leu(301), and alpha(1)Glu(303) only reacted with pCMBS(-) applied in the presence of GABA. We infer that the pCMBS(-) reactive residues are on the water-accessible surface of the protein and that GABA induces a conformational change that increases the water accessibility of the four M3 residues, possibly by inducing the formation of water-filled crevices that extend into the interior of the protein. Others have shown that mutations of alpha(1)Ala(291), a water-accessible residue, alter volatile anesthetic and ethanol potentiation of GABA-induced currents. Water-filled crevices penetrating into the interior of the membrane-spanning domain may allow anesthetics and alcohol to reach their binding sites and thus may have implications for the mechanisms of action of these agents.  (+info)

Cysteine residues in the Na+/dicarboxylate co-transporter, NaDC-1. (7/190)

The role of cysteine residues in the Na(+)/dicarboxylate co-transporter (NaDC-1) was tested using site-directed mutagenesis. The transport activity of NaDC-1 was not affected by mutagenesis of any of the 11 cysteine residues, indicating that no individual cysteine residue is necessary for function. NaDC-1 is sensitive to inhibition by the impermeant cysteine-specific reagent, p-chloromercuribenzenesulphonate (pCMBS). The pCMBS-sensitive residues in NaDC-1 are Cys-227, found in transmembrane domain 5, and Cys-476, located in transmembrane domain 9. Although cysteine residues are not required for function in NaDC-1, their presence appears to be important for protein stability or trafficking to the plasma membrane. There was a direct relationship between the number of cysteine residues, regardless of location, and the transport activity and expression of NaDC-1. The results indicate that mutagenesis of multiple cysteine residues in NaDC-1 may alter the shape or configuration of the protein, leading to alterations in protein trafficking or stability.  (+info)

Requirement of aquaporin-1 for NaCl-driven water transport across descending vasa recta. (8/190)

Deletion of AQP1 in mice results in diminished urinary concentrating ability, possibly related to reduced NaCl- and urea gradient-driven water transport across the outer medullary descending vasa recta (OMDVR). To quantify the role of AQP1 in OMDVR water transport, we measured osmotically driven water permeability in vitro in microperfused OMDVR from wild-type, AQP1 heterozygous, and AQP1 knockout mice. OMDVR diameters in AQP1(-/-) mice were 1.9-fold greater than in AQP1(+/+) mice. Osmotic water permeability (P(f)) in response to a 200 mM NaCl gradient (bath > lumen) was reduced about 2-fold in AQP1(+/-) mice and by more than 50-fold in AQP1(-/-) mice. P(f) increased from 1015 to 2527 microm/s in AQP1(+/+) mice and from 22 to 1104 microm/s in AQP1(-/-) mice when a raffinose rather than an NaCl gradient was used. This information, together with p-chloromercuribenzenesulfonate inhibition measurements, suggests that nearly all NaCl-driven water transport occurs by a transcellular route through AQP1, whereas raffinose-driven water transport also involves a parallel, AQP1-independent, mercurial-insensitive pathway. Interestingly, urea was also able to drive water movement across the AQP1-independent pathway. Diffusional permeabilities to small hydrophilic solutes were comparable in AQP1(+/+) and AQP1(-/-) mice but higher than those previously measured in rats. In a mathematical model of the medullary microcirculation, deletion of AQP1 resulted in diminished concentrating ability due to enhancement of medullary blood flow, partially accounting for the observed urine-concentrating defect.  (+info)

4-Chloromercuribenzenesulfonate is a chemical compound with the formula C6H5ClHgSO3. It is an organomercury compound, where mercury is bonded to a phenyl ring and a sulfonate group. This compound is an white crystalline powder that is soluble in water and denser than water.

It has been used historically as a diuretic and antiseptic, but its use in medicine has been discontinued due to the toxicity of mercury. Exposure to mercury can have serious health consequences, including damage to the nervous system, kidneys, and digestive system. Therefore, handling and disposal of 4-chloromercuribenzenesulfonate should be done with caution and in accordance with local regulations for hazardous materials.

4,4'-(3-oxo-1,5-pentanediyl)bis(n,n-dimethyl-n-2-propenyl-), dibromide MeSH D02.092.146.325 - p-dimethylaminoazobenzene MeSH ... 4-nitrophenyl) ester MeSH D02.705.539.783 - phorate MeSH D02.705.539.790 - phosmet MeSH D02.705.539.900 - temefos MeSH D02.705. ... 4-nitrophenyl) ester MeSH D02.886.309.783 - phorate MeSH D02.886.309.790 - phosmet MeSH D02.886.309.900 - temefos MeSH D02.886. ... 4,5-trisphosphate MeSH D02.033.800.519.400.700 - phytic acid MeSH D02.033.800.609 - mannitol MeSH D02.033.800.609.450 - ...
Benga G, Popescu O, Pop VI, Holmes RP (1986). "p-(Chloromercuri)benzenesulfonate binding by membrane proteins and the ... 265 (4 Pt 2): F463-76. doi:10.1152/ajprenal.1993.265.4.F463. PMID 7694481. S2CID 2685263. Mitsuoka K, Murata K, Walz T, Hirai T ... 126 (4): 1358-1369. doi:10.1104/pp.126.4.1358. PMC 117137. PMID 11500536. Sade, N; Shatil-Cohen, A; Attia, Z; Maurel, C; ... 81 (4): 361-8. doi:10.1016/S0888-7543(03)00029-6. PMID 12676560. Radin, M. Judith; Yu, Ming-Jiun; Stoedkilde, Lene; Miller, R ...
Benga, Gheorghe; Popescu, Octavian; Pop, Victor I.; Holmes, Ross P. (1986). "p-(Chloromercuri)benzenesulfonate binding by ... 1998 (4). Archived from the original on 14 March 2008. Kehe, Marjorie (2 October 2008). "Are US Writers Unworthy of the Nobel ... 50 (3-4): 167-172. PMID 20055226. Cox, Francis E. G. (2010). "History of the discovery of the malaria parasites and their ... Retrieved 4 January 2009.{{cite web}}: CS1 maint: unfit URL (link) "Textos escondidos de Pablo Neruda". Libros. 14 April 2005. ...
4,4-(3-oxo-1,5-pentanediyl)bis(n,n-dimethyl-n-2-propenyl-), dibromide MeSH D02.092.146.325 - p-dimethylaminoazobenzene MeSH ... 4-nitrophenyl) ester MeSH D02.705.539.783 - phorate MeSH D02.705.539.790 - phosmet MeSH D02.705.539.900 - temefos MeSH D02.705. ... 4-nitrophenyl) ester MeSH D02.886.309.783 - phorate MeSH D02.886.309.790 - phosmet MeSH D02.886.309.900 - temefos MeSH D02.886. ... 4,5-trisphosphate MeSH D02.033.800.519.400.700 - phytic acid MeSH D02.033.800.609 - mannitol MeSH D02.033.800.609.450 - ...
Y1 - 1988/4/13. N2 - The effects of ADP, ATP, 2-methylthio ATP, α,β-methylene ADP (α,βmeADP) and α,βmeATP on smooth muscle tone ...
4-Chloromercuribenzenesulfonate D1.632.750.740.220 5-Lipoxygenase-Activating Proteins D12.644.360.24.32 D12.776.157.57.00 ...
4-Chloromercuribenzenesulfonate D1.632.750.740.220 5-Lipoxygenase-Activating Proteins D12.644.360.24.32 D12.776.157.57.00 ...
4 Hydroxyphenylpyruvate Dioxygenase Deficiency Disease use Tyrosinemias 4-Nitrophenol-2-Hydroxylase use Cytochrome P-450 CYP2E1 ... 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer ... 2H-Benzo(a)quinolizin-2-ol, 2-Ethyl-1,3,4,6,7,11b-hexahydro-3-isobutyl-9,10-dimethoxy- ... 3-Keto-5-alpha-Steroid delta-4-Dehydrogenase use Testosterone 5-alpha-Reductase ...
1990 Oct;259(4 Pt 2):F539-44. doi: 10.1152/ajprenal.1990.259.4.F539. Am J Physiol. 1990. PMID: 2171356 ...
4-Chloromercuribenzenesulfonate D1.632.750.740.220 5-Lipoxygenase-Activating Proteins D12.644.360.24.32 D12.776.157.57.00 ...
CHLOROMERCURIBENZENESULFONATE 1975-93. History Note:. 94; was CHLOROMERCURIPHENYLSULFONATE (see under PHENYLMERCURY COMPOUNDS ... 4-Chloromercuribenzenesulfonate - Preferred Concept UI. M0004172. Scope note. A cytotoxic sulfhydryl reagent that inhibits ... 4-Chloromercurio-benzènesulfonate Entry term(s):. 4 Chloromercuribenzenesulfonate. Acid, Chloromercuribenzene-p-sulphonic. ... Mercurate(1-), chloro(4-sulfonatophenyl)-, hydrogen Previous Indexing:. Benzene (1966-1967). Benzenesulfonates (1974). Mercury ...
Chloromercuribenzenesulfonate Term UI T007917. Date05/10/1993. LexicalTag NON. ThesaurusID NLM (1994). ... Chloromercuribenzenesulfonate Chloromercuriphenylsulfonate PCMBS PCMPS p-Chloromercuriphenylsulphonate Pharm Action. Enzyme ... CHLOROMERCURIBENZENESULFONATE was see CHLOROMERCURIPHENYLSULFONATE 1975-93. Date Established. 1991/01/01. Date of Entry. 1974/ ... CHLOROMERCURIBENZENESULFONATE 1975-93. History Note. 94; was CHLOROMERCURIPHENYLSULFONATE (see under PHENYLMERCURY COMPOUNDS ...
Chloromercuribenzenesulfonate Term UI T007917. Date05/10/1993. LexicalTag NON. ThesaurusID NLM (1994). ... Chloromercuribenzenesulfonate Chloromercuriphenylsulfonate PCMBS PCMPS p-Chloromercuriphenylsulphonate Pharm Action. Enzyme ... CHLOROMERCURIBENZENESULFONATE was see CHLOROMERCURIPHENYLSULFONATE 1975-93. Date Established. 1991/01/01. Date of Entry. 1974/ ... CHLOROMERCURIBENZENESULFONATE 1975-93. History Note. 94; was CHLOROMERCURIPHENYLSULFONATE (see under PHENYLMERCURY COMPOUNDS ...
27 N0000170401 D-Ala(2),MePhe(4),Met(0)-ol-enkephalin N0000168380 D-Alanine Transaminase N0000167841 D-Amino-Acid Oxidase ... 4,4-(3-oxo-1,5-pentanediyl)bis(N,N-dimethyl-N-2-propenyl-), Dibromide N0000166410 Benzene N0000007525 Benzene Derivatives ... 4,5-Tetrahydro-8-chloro-3-methyl-5-phenyl-1H-3-benzazepin-7-ol N0000179007 1 alpha-hydroxyergocalciferol N0000179008 1,1,1- ... 4-alpha-Glucosidase N0000167706 Glucan 1,4-beta-Glucosidase N0000167709 Glucan Endo-1,3-beta-D-Glucosidase N0000007623 Glucans ...
Type 4 D8.811.913.696.620.682.725.400.46 D8.811.913.696.620.682.725.400.180 D12.776.543.750.60.116 D12.776.543.750.60.443 ... Interleukin-4 D23.50.301.264.35.705 D23.101.100.110.705 Receptors, Interleukin-6 D23.50.301.264.35.710 D23.101.100.110.710 ... D3.383.742.680.852.250 4-Chloromercuribenzenesulfonate D2.691.750.740.250 D2.691.750.740.220 5-Guanylic Acid D3.438.759.646. ... D23.529.624.130 Fibroblast Growth Factor 4 D12.644.276.750.140 D12.644.276.624.140 D23.348.383.140 D12.776.467.624.140 D23.348. ...
4-Chloromercuribenzenesulfonate D1.632.750.740.220 5-Lipoxygenase-Activating Proteins D12.644.360.24.32 D12.776.157.57.00 ...
... or by reacting with 60 mol of p-chloromercuribenzene sulfonate/10(6) g of protein. The effects of these activating procedures ... 4. Rats placed 0.65 m below a 250 W infra-red lamp responded to the imposed heat load by vasodilation of the tail skin blood ... 4. In most tissues and animals investigated, the values of the Km of adenosine kinase for adenosine are between one and two ... In dipeptides, SO-4 produces decarboxylation of the amino acid located at the carboxylate terminal residue. For gly-ala and ala ...

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