A mechanistic investigation of the thiol-disulfide exchange step in the reductive dehalogenation catalyzed by tetrachlorohydroquinone dehalogenase

Tetrachlorohydroquinone dehalogenase catalyzes the reductive dehalogenation of tetrachloro- and trichlorohydroquinone to give 2,6-dichlorohydroquinone in the pathway for degradation of pentachlorophenol by Sphingobium chlorophenolicum. Previous work has suggested that this enzyme may have originated from a glutathione-dependent double bond isomerase such as maleylacetoacetate isomerase or maleylpyruvate isomerase. While some of the elementary steps in these two reactions may be similar, the final step in the dehalogenation reaction, a thiol-disulfide exchange reaction that removes glutathione covalently bound to Cys13, certainly has no counterpart in the isomerization reaction. The thiol-disulfide exchange reaction does not appear to have been evolutionarily optimized. There is little specificity for the thiol; many thiols react at high rates. TCHQ dehalogenase binds the glutathione involved in the thioldisulfide exchange reaction very poorly and does not alter its pK(a) in order to improve its nucleophilicity. Remarkably, single-turnover kinetic studies show that the enzyme catalyzes this step by approximately 10000-fold. This high reactivity requires an as yet unidentified protonated group in the active site.