22752-72-3Relevant articles and documents
Substrate specificity of Sphingobium chlorophenolicum 2,6- dichlorohydroquinone 1,2-dioxygenase
MacHonkin, Timothy E.,Doerner, Amy E.
scheme or table, p. 8899 - 8913 (2012/05/05)
PcpA is an aromatic ring-cleaving dioxygenase that is homologous to the well-characterized Fe(II)-dependent catechol extradiol dioxygenases. This enzyme catalyzes the oxidative cleavage of 2,6-dichlorohydroquinone in the catabolism of pentachlorophenol by Sphingobium chlorophenolicum ATCC 39723. 1H NMR and steady-state kinetics were used to determine the regiospecificity of ring cleavage and the substrate specificity of the enzyme. PcpA exhibits a high degree of substrate specificity for 2,6-disubstituted hydroquinones, with halogens greatly preferred at those positions. Notably, the kcat app/KmAapp of 2,6-dichlorohydroquinone is ～40-fold higher than that of 2,6-dimethylhydroquinone. The asymmetric substrate 2-chloro-6-methylhydroquinone yields a mixture of 1,2- and 1,6-cleavage products. These two modes of cleavage have different K mO2app values (21 and 260 μM, respectively), consistent with a mechanism in which the substrate binds in two catalytically productive orientations. In contrast, monosubstituted hydroquinones show a limited amount of ring cleavage but rapidly inactivate the enzyme in an O2-dependent fashion, suggesting that oxidation of the Fe(II) may be the cause. Potent inhibitors of PcpA include ortho-disubstituted phenols and 3-bromocatechol. 2,6-Dibromophenol is the strongest competitive inhibitor, consistent with PcpA's substrate specificity. Several factors that could yield this specificity for halogen substituents are discussed. Interestingly, 3-bromocatechol also inactivates the enzyme, while 2,6-dihalophenols do not, indicating a requirement for two hydroxyl groups for ring cleavage and for enzyme inactivation. These results provide mechanistic insights into the hydroquinone dioxygenases.
