Purification and Characterization of meta-Cleavage Compound Hydrolase from a Carbazole Degrader Pseudomonas resinovorans Strain CA10(Microbiology & Fermentation Technology)

概要

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2-Hydroxy-6-oxo-6-(2'-aminophenyl)-hexa-2,4-dienoic acid [6-(2'-aminophenyl)-HODA] hydrolase, involved in carbazole degradation by Pseudomonas resinovorans strain CA10, was purified to near homogeneity from an overexpressing Escherichia coli strain. The enzyme was dimeric, and its optimum pH was 7.0-7.5. Phylogenetic analysis showed the close relationship of this enzyme to other hydrolases involved in the degradation of monocyclic aromatic compounds, and this enzyme was specific for 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (6-phenyl-HODA), having little activity toward 2-hydroxy-6-oxohepta-2,4-dienoic acid and 2-hydroxymuconic semialdehyde. The enzyme had a K_m of 2.51 μΜ and K_<cat> of 2.14 (s^<-1) for 6-phenyl-HODA (50 mM sodium phosphate, pH 7.5, 25℃). The effect of the presence of an amino group or hydroxyl group at the 2'-position of phenyl moiety of 6-phenyl-HODA on the enzyme activity was found to be small ; the activity decreased only in the order of 6-(2'-aminophenyl)-HODA (2.44 U/mg)> 6-phenyl-HODA (1.99U / mg)>2-hydroxy-6-oxo-6-(2'-hydroxyphenyl)-hexa-2,4-dienoic acid (1.05 U/mg). The effects of 2'-substitution on the activity were in accordance with the predicted reactivity based on the calculated lowest unoccupied molecular orbital energy for these substrates.
社団法人日本農芸化学会の論文
2003-01-23