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Applied and Environmental Microbiology, June 2007, p. 3575-3580, Vol. 73, No. 11
0099-2240/07/$08.00+0     doi:10.1128/AEM.00011-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Characterization of SafC, a Catechol 4-O-Methyltransferase Involved in Saframycin Biosynthesis{triangledown}

James T. Nelson,1,{dagger} Jaeheon Lee,2,{dagger} James W. Sims,1 and Eric W. Schmidt1,2*

Department of Medicinal Chemistry,1 Department of Biology, University of Utah, Salt Lake City, Utah 841122

Received 3 January 2007/ Accepted 10 April 2007

Members of the saframycin/safracin/ecteinascidin family of peptide natural products are potent antitumor agents currently under clinical development. Saframycin MX1, from Myxococcus xanthus, is synthesized by a nonribosomal peptide synthetase, SafAB, and an O-methyltransferase, SafC, although other proteins are likely involved in the pathway. SafC was overexpressed in Escherichia coli, purified to homogeneity, and assayed for its ability to methylate a variety of substrates. SafC was able to catalyze the O-methylation of catechol derivatives but not phenols. Among the substrates tested, the best substrate for SafC was L-dihydroxyphenylalanine (L-dopa), which was methylated specifically in the 4'-O position (kcat/Km = 5.5 x 103 M–1 s–1). SafC displayed less activity on other catechol derivatives, including catechol, dopamine, and caffeic acid. The more labile L-5'-methyldopa was an extremely poor substrate for SafC (kcat/Km = ~2.8 x 10–5 M–1 s–1). L-Dopa thioester derivatives were also much less reactive than L-dopa. These results indicate that SafC-catalyzed 4'-O-methylation of L-dopa occurs prior to 5'-C-methylation, suggesting that 4'-O-methylation is likely the first committed step in the biosynthesis of saframycin MX1. SafC has biotechnological potential as a methyltransferase with unique regioselectivity.

* Corresponding author. Mailing address: Department of Medicinal Chemistry, University of Utah, 30 South 2000 East, Room 201, Salt Lake City, UT 84112. Phone: (801) 585-5234. Fax: (801) 585-9119. E-mail: ews1{at}utah.edu

{triangledown} Published ahead of print on 20 April 2007.

{dagger} J.T.N. and J.L. contributed equally to this work.

Applied and Environmental Microbiology, June 2007, p. 3575-3580, Vol. 73, No. 11
0099-2240/07/$08.00+0     doi:10.1128/AEM.00011-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Kopycki, J. G., Stubbs, M. T., Brandt, W., Hagemann, M., Porzel, A., Schmidt, J., Schliemann, W., Zenk, M. H., Vogt, T. (2008). Functional and Structural Characterization of a Cation-dependent O-Methyltransferase from the Cyanobacterium Synechocystis sp. Strain PCC 6803. J. Biol. Chem. 283: 20888-20896 [Abstract] [Full Text]  


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