Mechanism of action of the redox affinity reagent [(trimethylammonio)methyl]catechol.
The synthesis of 4- and 5-hydroxy-3-[(trimethylammonio)methyl]catechol (4- and 5-HTMC) was carried out to examine their proposed involvement as intermediates in the spontaneous redox-dependent half-of-sites inactivation of neurotoxin binding sites in the nicotinic acetylcholine receptor (nAcChR) mediated by the parent compound 3-[(trimethylammonio)methyl]catechol (TMC) [Nickoloff et al. (1985) Biochemistry 24, 999-1007]. Oxidation of 4- and 5-HTMC occurred with sodium periodate with facile conversion to the corresponding p-quinones which were intercepted with thiols and cyclopentadiene. Both 4- and 5-HTMC inactivated neurotoxin binding in the nAcChR in a time course and over a concentration range consistent with their involvement as intermediates in the TMC redox-dependent inactivation of neurotoxin ([125I]-alpha-bungarotoxin) binding sites. Rapid concentration-dependent inactivation of neurotoxin sites occurred over a 10-1000 microM range and was resistant to further inactivation after 50% loss of available toxin binding sites on the nAcChR. Both 4- and 5-HTMC inactivated nAcChR neurotoxin sites much more rapidly and efficiently than was observed previously with TMC. The apparent binding constants for 4- and 5-HTMC with the nAcChR, calculated from their concentration-dependent inactivation behavior toward toxin binding sites, were Kd = 224 +/- 98 and 39 +/- 17 microM, respectively. The observed results and the redox potentials (vs Ag/AgCl reference electrode) measured by cyclic voltammetry at pH 1.8 for TMC (719 mV) and the 4- and 5-HTMC derivatives (519 and 443 mV, respectively) supported the previously proposed mechanism for inactivation of the nAcChR by TMC.(ABSTRACT TRUNCATED AT 250 WORDS)
Gu
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Department of Medicinal and Biological Chemistry, College of Pharmacy, University of Toledo, Ohio 43606.
Lee
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Kirchhoff
J R
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Manzey
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Hudson
R A
RA
D001548:Benzalkonium Compounds / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites; D002038:Bungarotoxins / Q000378:metabolism; D002396:Catechols / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D004229:Dithiothreitol / Q000494:pharmacology; D004563:Electrochemistry; D007202:Indicators and Reagents; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010084:Oxidation-Reduction; D011809:Quinones / Q000378:metabolism; D011978:Receptors, Nicotinic / Q000187:drug effects* / Q000378:metabolism; D013053:Spectrophotometry; D013816:Thermodynamics
D001548:Benzalkonium Compounds; D002038:Bungarotoxins; D002396:Catechols; D007202:Indicators and Reagents; D011809:Quinones; D011978:Receptors, Nicotinic; C088430:4-hydroxy-3-((trimethylammonio)methyl)catechol; C088431:5-hydroxy-3-((trimethylammonio)methyl)catechol; C036022:3-trimethylaminomethylcatechol; D004229:Dithiothreitol