Oxidative Inactivation of Ring-Cleavage Extradiol Dioxygenases: Mechanism and Ferredoxin-Mediated Reactivation
Extradiol dioxygenases are ubiquitous enzymes that catalyze ring cleavage of a wide variety of aromatic compounds. Most of these enzymes contain a ferrous ion at the active site, which is bound to the polypeptide chain through a conserved triad of two his
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K. N. Timmis (ed.), Handbook of Hydrocarbon and Lipid Microbiology, DOI 10.1007/978-3-540-77587-4_75, # Springer-Verlag Berlin Heidelberg, 2010
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Oxidative Inactivation of Ring-Cleavage Extradiol Dioxygenases
Abstract: Extradiol dioxygenases are ubiquitous enzymes that catalyze ring cleavage of a wide variety of aromatic compounds. Most of these enzymes contain a ferrous ion at the active site, which is bound to the polypeptide chain through a conserved triad of two histidines and one glutamic acid. During the catalytic cycle, a catecholic substrate first binds at the active site, followed by dioxygen and the ternary complex formed yields a bound superoxide that attacks the substrate, leading eventually to ring cleavage. The active site iron remains ferrous during catalysis, except when poor substrates such as chloro- or methylcatechols are processed. In such cases, the enzyme becomes inactivated through oxidation and eventually loss of its active site iron atom. In Pseudomonas putida mt-2, catechol 2,3-dioxygenase, which is involved in toluene degradation, is inactivated by 4-methylcatechol. The enzyme is however rescued by a specific reactivation system involving a [2Fe-2S] ferredoxin encoded by xylT. The role of this ferredoxin is to reduce the ferric ion of the inactive enzyme thereby regenerating the active catalyst. Recent findings indicate that the electrons needed for the XylT-mediated reactivation are provided by XylZ, a NADH-oxidoreductase that is part of the toluate dioxygenase complex. XylT analogues present in other bacteria have been shown to have a similar role in the reactivation of catechol dioxygenases involved in the degradation of various aromatic hydrocarbons including cresols, chlorobenzene and naphthalene. The occurrence and significance of ferredoxin-mediated extradiol dioxygenae repair systems in bacteria is discussed.
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Introduction
The cleavage of the aromatic ring is a critical step in the biodegradation of aromatic hydrocarbons by aerobic bacteria. It is catalyzed by two kinds of enzyme, the intradiol and extradiol dioxygenases, which represent two classes of phylogenetically unrelated proteins (Harayama et al., 1992). Although both types of enzymes are active on similar catecholic substrates they have different structures and catalytic mechanisms (Vaillancourt et al., 2006). Also, extradiol dioxygenases appear to be more versatile than intradiol dioxygenases as they react with a wider range of substrates and intervene in a greater variety of pathways. However, they are labile enzymes which undergo oxidative inactivation, especially in the presence of poor substrates such as alkyl- or chloro-substituted catechols. Current knowledge of the mechanism underlying extradiol dioxygenase inactivation is reviewed here, in the light of recent reports describing in detail the catalytic cycle of this type of enzyme and the process of mechanism-based enzyme inactivation. A natural repair system, preventing inactivation of catechol 2,3-dioxygenase, was first discovered in Pseudomonas p
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