Molecular basis for the substrate selectivity of bicyclic and monocyclic extradiol dioxygenases

Vaillancourt, Frédéric H.; Fortin, Pascal D.; Labbé, Geneviève; Drouin, Nathalie M.; Karim, Zamil; Agar, Nathalie Y. R.; Eltis, Lindsay D.

doi:10.1016/j.bbrc.2005.08.219

Cited by 10 publications

(11 citation statements)

References 33 publications

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“…5 The structural homogeneity of our substrate suite further emphasizes this conclusion. Second, the K M for oxygen is an essential component of these interpretations, and others have made the same observations; however, in these cases, catechol-2,3-dioxygenase 12 and DHBD, 15 the K M for oxygen is very high for the "native" substrate, so high that oxygen cannot be saturating under conditions where the catecholic substrate is varied. In the case of the VOC enzyme catechol 2,3-dioxygenase, the authors measure the second-order rate for oxygen (k cat /K M O 2 that is equivalent to a k SP for oxygen) and conclude that increasing the electron-withdrawing nature of the substituent compromises oxygen binding and catalysis.…”

Section: ■ Conclusionmentioning

confidence: 90%

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

et al. 2021

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Dioxygenase enzymes are essential protein catalysts for the breakdown of catecholic rings, structural components of plant woody tissue. This powerful chemistry is used in nature to make antibiotics and other bioactive materials or degrade plant material, but we have a limited understanding of the breadth and depth of substrate space for these potent catalysts. Here we report steady-state and pre-steady-state kinetic analysis of dopamine derivatives substituted at the 6-position as substrates of L-DOPA dioxygenase, and an analysis of that activity as a function of the electron-withdrawing nature of the substituent. Steady-state and pre-steady-state kinetic data demonstrate the dopamines are impaired in binding and catalysis with respect to the cosubstrate molecular oxygen, which likely afforded spectroscopic observation of an early reaction intermediate, the semiquinone of dopamine. The reaction pathway of dopamine in the pre-steady state is consistent with a nonproductive mode of binding of oxygen at the active site. Despite these limitations, L-DOPA dioxygenase is capable of binding all of the dopamine derivatives and catalyzing multiple turnovers of ring cleavage for dopamine, 6-bromodopamine, 6-carboxydopamine, and 6-cyanodopamine. 6-Nitrodopamine was a single-turnover substrate. The variety of substrates accepted by the enzyme is consistent with an interplay of factors, including the capacity of the active site to bind large, negatively charged groups at the 6-position and the overall oxidizability of each catecholamine, and is indicative of the utility of extradiol cleavage in semisynthetic and bioremediation applications.

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Section: ■ Conclusionmentioning

confidence: 90%

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

et al. 2021

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“…Deveryshetty and Phale (57) demonstrated competitive inhibition of 1-hydroxy-2-naphthoic acid dioxygenase by a stereoisomeric compound, 3-hydroxy-2-naphthoic acid. An alternative mechanism, oxidative (suicide) inactivation of extradiol dioxygenases, has been demonstrated with suboptimal substrates, and the level of inhibition depended on the sequence variation of catabolic enzymes (68). Similar mechanisms may cause enzyme inhibition and concomitant diminished growth/ degradation if the alternative pathway intermediates of antagonized mixture catabolism interfere at enzyme active sites.…”

Section: Discussionmentioning

confidence: 99%

Effects of Polycyclic Aromatic Hydrocarbon Mixtures on Degradation, Gene Expression, and Metabolite Production in Four Mycobacterium Species

Hennessee

2016

Appl Environ Microbiol

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Polycyclic aromatic hydrocarbons (PAHs) are widespread environmental contaminants that are hazardous to human health. It has been demonstrated that members of the Mycobacterium genus are among the most effective degraders of PAHs, but few studies have focused on the degradation of PAH mixtures. In this study, single and mixed PAH metabolism was investigated in four phylogenetically distinct Mycobacterium species with respect to (i) parent compound degradation, (ii) bacterial growth, (iii) catabolic gene expression, and (iv) metabolite production. Synergistic and antagonistic effects on four model PAH compounds (benzo[a]pyrene, pyrene, fluoranthene, and phenanthrene) characterized degradation of mixtures in a strain-and mixture-dependent manner. The mixture of pyrene and phenanthrene, in particular, resulted in antagonized degradation by three out of four bacterial species, and further studies were narrowed to investigate the degradation of this mixture. Antagonistic effects persisted over time and were correlated with reduced bacterial growth. Antagonized degradation of PAH was not caused by preferential degradation of secondary PAHs, nor were mixture compounds or concentrations toxic to cells growing on sugars. Reverse transcription-PCR (RT-PCR) studies of the characterized catabolic pathway of phenanthrene showed that in one organism, antagonism of mixture degradation was associated with downregulated gene expression. Metabolite profiling revealed that antagonism in mixture degradation was associated with the shunting of substrate through alternative pathways not used during the degradation of single PAHs. The results of this study demonstrate metabolic differences between single and mixed PAH degradation with consequences for risk assessment and bioremediation of PAH-contaminated sites. IMPORTANCEMycobacterium species are promising organisms for environmental bioremediation because of their ubiquitous presence in soils and their ability to catabolize aromatic compounds. PAHs can be degraded effectively as single compounds, but mixed substrates often are subject to degradative inhibition, which may explain the persistence of these pollutants in soils. Single and mixed PAH degradation by diverse Mycobacterium species was compared, with associated bacterial growth, gene expression, and metabolite production. The results demonstrate that antagonism characterized degradation in a strain-and mixture-dependent manner. One strain that was versatile in its pathway use of single chemicals also efficiently degraded the mixture, whereas antagonism in other the strains was associated with altered metabolic profiles, indicating unusual pathway use. The impacts of this work on risk assessment and bioremediation modeling studies indicate the need to account for mixture-generated intermediates and to recognize mixture degradation as a property distinct from that of PAH substrate range. P olycyclic aromatic hydrocarbons (PAHs) are a class of chemical pollutants that are widespread and persistent in the environment (1-3...

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“…3). As in Vaillancourt's report (Vaillancourt et al , 2005), the active site of BphC was poorly complementary to catechol because the volume in the active site occupied by the second ring of biphenyl was excess ‘free’ volume when catechol binds. Therefore, the specificity of BphC for bicyclic substrate was higher than that for monocyclic substrate.…”

Section: Resultsmentioning

confidence: 86%

“…Comparing the 3D structures of BphC_LA‐4, BphC_LB400 and BphC_KKS102, the β‐strand between two α‐helices was absent in the second βαβββ motif of BphC_LA‐4 in the N‐terminal domain (Han et al , 1995; Senda et al , 1996). According to the enzyme–substrate complexes, the volume in the active site occupied by the second ring of biphenyl played an important role in the determination of the specificity of BphC for bicyclic and monocyclic catecholic substrates (Vaillancourt et al , 2005). As shown in Fig.…”

Section: Discussionmentioning

confidence: 99%

Enzymeâsubstrate interaction and characterization of a 2,3-dihydroxybiphenyl 1,2-dioxygenase fromDyella ginsengisoliLA-4

2009

FEMS Microbiology Letters

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A bphC gene (915 bp) encoding 2,3-dihydroxybiphenyl 1,2-dioxygenase (BphC) was amplified by PCR from Dyella ginsengisoli LA-4, which was heterologously expressed in Escherichia coli. The purified His-Tag BphC was able to catalyze the meta-cleavage reaction of the dihydroxylated aromatic rings. According to the specificity constant (K(cat)/K(m)) of BphC_LA-4, the specificity of BphC_LA-4 was determined in the following order: 2,3-dihydroxybiphenyl>3-methylcatechol>catechol>4-chlorocatechol>4-methylcatechol. The experimental data were consistent with the prediction of enzyme-substrate complexes. The highest specific activity of BphC_LA-4 was 118.3 U mg(-1) for 2,3-dihydroxybiphenyl.

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Molecular basis for the substrate selectivity of bicyclic and monocyclic extradiol dioxygenases

Cited by 10 publications

References 33 publications

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

Effects of Polycyclic Aromatic Hydrocarbon Mixtures on Degradation, Gene Expression, and Metabolite Production in Four Mycobacterium Species

Enzymeâsubstrate interaction and characterization of a 2,3-dihydroxybiphenyl 1,2-dioxygenase fromDyella ginsengisoliLA-4

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Molecular basis for the substrate selectivity of bicyclic and monocyclic extradiol dioxygenases

Cited by 10 publications

References 33 publications

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

L-DOPA Dioxygenase Activity on 6-Substituted Dopamine Analogues

Effects of Polycyclic Aromatic Hydrocarbon Mixtures on Degradation, Gene Expression, and Metabolite Production in Four Mycobacterium Species

Enzymeâsubstrate interaction and characterization of a 2,3-dihydroxybiphenyl 1,2-dioxygenase fromDyella ginsengisoliLA-4

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Enzymeâsubstrate interaction and characterization of a 2,3-dihydroxybiphenyl 1,2-dioxygenase fromDyella ginsengisoliLA-4