Identification of a Novel Self-Sufficient Styrene Monooxygenase from Rhodococcus opacus 1CP
| Autor: | Stefan R. Kaschabek, Willem J. H. van Berkel, Dirk Eulberg, Michael Schlömann, Silvia Lakner, Dirk Tischler |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2009 |
| Předmět: |
erythropolis
Molecular Sequence Data enzymes Flavoprotein Biochemie Flavin group Microbiology Models Biological Biochemistry Cofactor Gas Chromatography-Mass Spectrometry chemistry.chemical_compound Rhodococcus opacus Bacterial Proteins Styrene oxide Flavin reductase electrochemical regeneration Rhodococcus catabolic pathway Molecular Biology Chromatography High Pressure Liquid Phylogeny Styrene VLAG degradation Flavin adenine dinucleotide biology Monooxygenase Enzymes and Proteins cytochrome-p450 Recombinant Proteins chemistry coenzyme regeneration biology.protein Oxygenases escherichia-coli Epoxy Compounds multiple sequence alignment Genome Bacterial pseudomonas-fluorescens st |
| Zdroj: | Journal of Bacteriology, 191(15), 4996-5009 Journal of Bacteriology 191 (2009) 15 |
| ISSN: | 0021-9193 |
| Popis: | Sequence analysis of a 9-kb genomic fragment of the actinobacterium Rhodococcus opacus 1CP led to identification of an open reading frame encoding a novel fusion protein, StyA2B, with a putative function in styrene metabolism via styrene oxide and phenylacetic acid. Gene cluster analysis indicated that the highly related fusion proteins of Nocardia farcinica IFM10152 and Arthrobacter aurescens TC1 are involved in a similar physiological process. Whereas 413 amino acids of the N terminus of StyA2B are highly similar to those of the oxygenases of two-component styrene monooxygenases (SMOs) from pseudomonads, the residual 160 amino acids of the C terminus show significant homology to the flavin reductases of these systems. Cloning and functional expression of His10-StyA2B revealed for the first time that the fusion protein does in fact catalyze two separate reactions. Strictly NADH-dependent reduction of flavins and highly enantioselective oxygenation of styrene to (S)-styrene oxide were shown. Inhibition studies and photometric analysis of recombinant StyA2B indicated the absence of tightly bound heme and flavin cofactors in this self-sufficient monooxygenase. StyA2B oxygenates a spectrum of aromatic compounds similar to those of two-component SMOs. However, the specific activities of the flavin-reducing and styrene-oxidizing functions of StyA2B are one to two orders of magnitude lower than those of StyA/StyB from Pseudomonas sp. strain VLB120. The incorporation of one atom of oxygen during hydroxylation, epoxidation, sulfoxidation, or Baeyer-Villiger oxidation is a common initial step of the aerobic degradation of aromatic compounds by microorganisms. In bacteria, these reactions are most frequently catalyzed by inducible flavoprotein monooxygenases (EC 1.14.13 [57]). The majority of these enzymes (socalled single-component flavoprotein monooxygenases) utilize electrons from NAD(P)H, which are transferred to a noncovalently bound flavin adenine dinucleotide (FAD) in order to activate molecular oxygen as a flavin (hydro)peroxide. Depending on the protonation of this intermediate and the type of substrate, an oxygen atom is then incorporated by nucleophilic or electrophilic attack. More recently, different twocomponent flavoprotein monooxygenases have been characterized (57). These systems cover an NAD(P)H-dependent flavin reductase in order to generate reduced flavin and an oxygenase that utilizes this cofactor for the activation of oxygen. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|