title: Inhibition of carbonic anhydrase II by thioxolone: a mechanistic and structural study.
abstract: This paper examines the functional mechanism of thioxolone, a compound recently identified as a weak inhibitor of human carbonic anhydrase II by Iyer et al. (2006) J. Biomol. Screening 11, 782-791 . Thioxolone lacks sulfonamide, sulfamate, or hydroxamate functional groups that are typically found in therapeutic carbonic anhydrase (CA) inhibitors, such as acetazolamide. Analytical chemistry and biochemical methods were used to investigate the fate of thioxolone upon binding to CA II, including Michaelis-Menten kinetics of 4-nitrophenyl acetate esterase cleavage, liquid chromatography-mass spectrometry (LC-MS), oxygen-18 isotope exchange studies, and X-ray crystallography. Thioxolone is proposed to be a prodrug inhibitor that is cleaved via a CA II zinc-hydroxide mechanism known to catalyze the hydrolysis of esters. When thioxolone binds in the active site of CA II, it is cleaved and forms 4-mercaptobenzene-1,3-diol via the intermediate S-(2,4-thiophenyl)hydrogen thiocarbonate. The esterase cleavage product binds to the zinc active site via the thiol group and is therefore the active CA inhibitor, while the intermediate is located at the rim of the active-site cavity. The time-dependence of this inhibition reaction was investigated in detail. Because this type of prodrug inhibitor mechanism depends on cleavage of ester bonds, this class of inhibitors may have advantages over sulfonamides in determining isozyme specificity. A preliminary structure-activity relationship study with a series of structural analogues of thioxolone yielded similar estimates of inhibition constants for most compounds, although two compounds with bromine groups at the C1 carbon of thioxolone were not inhibitory, suggesting a possible steric effect.
authors.affiliation: Department of Biological Sciences, Mailstop 5410, College of Arts and Sciences, 1903 West Michigan Avenue, Western Michigan University, Kalamazoo, Michigan 49008-5410, USA.
authors.forename: Albert A
authors.initials: AA
authors.lastname: Barrese
authors.forename: Caroli
authors.initials: C
authors.lastname: Genis
authors.forename: S Zoe
authors.initials: SZ
authors.lastname: Fisher
authors.forename: Jared N
authors.initials: JN
authors.lastname: Orwenyo
authors.forename: Mudalige Thilak
authors.initials: MT
authors.lastname: Kumara
authors.forename: Subodh K
authors.initials: SK
authors.lastname: Dutta
authors.forename: Eric
authors.initials: E
authors.lastname: Phillips
authors.forename: James J
authors.initials: JJ
authors.lastname: Kiddle
authors.forename: Chingkuang
authors.initials: C
authors.lastname: Tu
authors.forename: David N
authors.initials: DN
authors.lastname: Silverman
authors.forename: Lakshmanan
authors.initials: L
authors.lastname: Govindasamy
authors.forename: Mavis
authors.initials: M
authors.lastname: Agbandje-McKenna
authors.forename: Robert
authors.initials: R
authors.lastname: McKenna
authors.forename: Brian C
authors.initials: BC
authors.lastname: Tripp
mesh_terms: D024402:Carbonic Anhydrase II / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002257:Carbonic Anhydrase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D002853:Chromatography, Liquid; D018360:Crystallography, X-Ray; D006801:Humans; D007700:Kinetics; D007783:Lactones / Q000737:chemistry / Q000494:pharmacology*; D013058:Mass Spectrometry; D013329:Structure-Activity Relationship
chemical_list: D002257:Carbonic Anhydrase Inhibitors; D007783:Lactones; C009474:thioxolone; D024402:Carbonic Anhydrase II