This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Kendrew, S. G. Articles by Marsh, E. N. Search for Related Content PubMed PubMed Citation Articles by Kendrew, S. G. Articles by Marsh, E. N.

 Previous Article  |  Next Article 

J. Bacteriol., Jul 1997, 4305-4310, Vol 179, No. 13
Copyright © 1997, American Society for Microbiology

Identification of a monooxygenase from Streptomyces coelicolor A3(2) involved in biosynthesis of actinorhodin: purification and characterization of the recombinant enzyme

SG Kendrew, DA Hopwood and EN Marsh
Department of Biochemistry and Cambridge Centre for Molecular Recognition, University of Cambridge, United Kingdom.

The oxidation of phenols to quinones is an important reaction in the oxidative tailoring of many aromatic polyketides from bacterial and fungal systems. Sequence similarity between ActVA-Orf6 protein from the actinorhodin biosynthetic cluster and the previously characterized TcmH protein that is involved in tetracenomycin biosynthesis suggested that ActVA-Orf6 might catalyze this transformation as a step in actinorhodin biosynthesis. To investigate the role of ActVA-Orf6 in this oxidation, we have expressed the actVA-Orf6 gene in Escherichia coli and purified and characterized the recombinant protein. ActVA-Orf6 was shown to catalyze the monooxygenation of the tetracenomycin intermediate TcmF1 to TcmD3, strongly suggesting that it catalyzes oxidation of a similar intermediate in actinorhodin biosynthesis. The monooxygenase obeys simple reaction kinetics and has a Km of 4.8 +/- 0.9 microM, close to the figure reported for the homologous enzyme TcmH. The enzyme contains no prosthetic groups and requires only molecular oxygen to catalyze the oxidation. Site-directed mutagenesis was used to investigate the role of histidine residues thought to be important in the reaction; mutants lacking His-52 displayed much-reduced activity, consistent with the proposed mechanistic hypothesis that this histidine acts as a general base during catalysis.

This article has been cited by other articles:

• Bab-Dinitz, E., Shmuely, H., Maupin-Furlow, J., Eichler, J., Shaanan, B. (2006). Haloferax volcanii PitA: an example of functional interaction between the Pfam chlorite dismutase and antibiotic biosynthesis monooxygenase families?. Bioinformatics 22: 671-675 [Abstract] [Full Text]  
• Chen, Y.-H., Wang, C.-C., Greenwell, L., Rix, U., Hoffmeister, D., Vining, L. C., Rohr, J., Yang, K.-Q. (2005). Functional Analyses of Oxygenases in Jadomycin Biosynthesis and Identification of JadH as a Bifunctional Oxygenase/Dehydrase. J. Biol. Chem. 280: 22508-22514 [Abstract] [Full Text]  
• Adams, M. A., Jia, Z. (2005). Structural and Biochemical Evidence for an Enzymatic Quinone Redox Cycle in Escherichia coli: IDENTIFICATION OF A NOVEL QUINOL MONOOXYGENASE. J. Biol. Chem. 280: 8358-8363 [Abstract] [Full Text]  
• Valton, J., Filisetti, L., Fontecave, M., Niviere, V. (2004). A Two-component Flavin-dependent Monooxygenase Involved in Actinorhodin Biosynthesis in Streptomyces coelicolor. J. Biol. Chem. 279: 44362-44369 [Abstract] [Full Text]  
• Ichinose, K., Ozawa, M., Itou, K., Kunieda, K., Ebizuka, Y. (2003). Cloning, sequencing and heterologous expression of the medermycin biosynthetic gene cluster of Streptomyces sp. AM-7161: towards comparative analysis of the benzoisochromanequinone gene clusters. Microbiology 149: 1633-1645 [Abstract] [Full Text]  
• Chung, J.-y., Fujii, I., Harada, S., Sankawa, U., Ebizuka, Y. (2002). Expression, Purification, and Characterization of AknX Anthrone Oxygenase, Which Is Involved in Aklavinone Biosynthesis in Streptomyces galilaeus. J. Bacteriol. 184: 6115-6122 [Abstract] [Full Text]