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Applied and Environmental Microbiology, June 2009, p. 3469-3476, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02590-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Application of a Newly Identified and Characterized 18-O-Acyltransferase in Chemoenzymatic Synthesis of Selected Natural and Nonnatural Bioactive Derivatives of Phoslactomycins{triangledown}

Mohini S. Ghatge, Nadaraj Palaniappan, Ma'moun M. Alhamadsheh, Jessica DiBari, and Kevin A. Reynolds*

Department of Chemistry, Portland State University, Portland, Oregon 97201

Received 12 November 2008/ Accepted 11 March 2009

Phoslactomycins (PLMs) and related leustroducsins (LSNs) have been isolated from a variety of bacteria based on antifungal, anticancer, and other biological assays. Streptomyces sp. strain HK 803 produces five PLM analogs (PLM A and PLMs C to F) in which the C-18 hydroxyl substituent is esterified with a range of branched, short-alkyl-chain carboxylic acids. The proposed pathway intermediate, PLM G, in which the hydroxyl residue is not esterified has not been observed at any significant level in fermentation, and the only route to this potentially useful intermediate has been an enzymatic deacylation of other PLMs and LSNs. We report that deletion of plmS3 from the PLM biosynthetic cluster gives rise to a mutant which accumulates the PLM G intermediate. The 921-bp plmS3 open reading frame was cloned and expressed as an N-terminally polyhistidine-tagged protein in Escherichia coli and shown to be an 18-O acyltransferase, catalyzing conversion of PLM G to PLM A, PLM C, and PLM E using isobutyryl coenzyme A (CoA), 3-methylbutyryl-CoA, and cyclohexylcarbonyl-CoA, respectively. The efficiency of this process (kcat of 28 ± 3 min–1 and Km of 88 ± 16 µM) represents a one-step chemoenzymatic alternative to a multistep synthetic process for selective chemical esterification of the C-18 hydroxy residue of PLM G. PlmS3 was shown to catalyze esterification of PLM G with CoA and N-acetylcysteamine thioesters of various saturated, unsaturated, and aromatic carboxylic acids and thus also to provide an efficient chemoenzymatic route to new PLM analogs.

* Corresponding author. Mailing address; Department of Chemistry, Portland State University, 262 Science Building 2, 1719 SW 10th Avenue, Portland, OR 97201. Phone: (503) 725-3886. Fax: (503) 725-9525. E-mail: reynoldsk{at}pdx.edu

{triangledown} Published ahead of print on 20 March 2009.

Applied and Environmental Microbiology, June 2009, p. 3469-3476, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02590-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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