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Two-component flavin-dependent pyrrole-2-carboxylate monooxygenase from Rhodococcus sp
D Becker1, T Schräder, J R Andreesen
1Institut für Mikrobiologie, Martin-Luther-Universität Halle, Germany.
European Journal of Biochemistry
|December 12, 1997
Summary
Rhodococcus sp. utilizes pyrrole-2-carboxylate for growth. A novel two-component flavin aromatic monooxygenase, comprising reductase and oxygenase proteins, catalyzes its degradation via NADH-dependent hydroxylation.
Area of Science:
- Biochemistry
- Microbiology
Background:
- Pyrrole-2-carboxylate serves as a sole carbon, nitrogen, and energy source for Rhodococcus sp.
- NADH-dependent oxygenase activity initiates substrate degradation.
Purpose of the Study:
- To purify and characterize the enzyme responsible for pyrrole-2-carboxylate degradation.
- To elucidate the mechanism and classification of this novel enzyme system.
Main Methods:
- Enzyme purification via chromatography.
- Enzymatic activity assays (NADH-dependent reduction of cytochrome c, NADH-oxidase activity, substrate-dependent oxygen consumption).
- Protein sequencing (proteolytic digestion and peptide sequencing).
Main Results:
- Two homogeneous protein components were isolated: an 18.7-kDa reductase and a 54-kDa oxygenase.
- The reductase component exhibited NADH-dependent activity, while the oxygenase required the reductase, FAD, and NADH for pyrrole-2-carboxylate hydroxylation.
- Sequence analysis revealed similarities to known two-component flavin aromatic monooxygenases, classifying the enzyme as such.
Conclusions:
- The pyrrole-2-carboxylate degradation is catalyzed by a novel two-component flavin aromatic monooxygenase from Rhodococcus sp.
- The interaction and ratio of the reductase and oxygenase components are crucial for efficient catalysis and suppression of uncoupled NADH oxidation.