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Antimicrobial Agents and Chemotherapy, July 1999, p. 1565-1573, Vol. 43, No. 7
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Identification and Analysis of the Balhimycin Biosynthetic Gene Cluster and Its Use for Manipulating Glycopeptide Biosynthesis in Amycolatopsis mediterranei DSM5908

S. Pelzer,1 R. Süßmuth,2 D. Heckmann,1,dagger J. Recktenwald,1 P. Huber,1 G. Jung,2 and W. Wohlleben1,*

Mikrobiologie/Biotechnologie1 and Institut für Organische Chemie,2 Eberhard-Karls-Universität Tübingen, D-72076 Tübingen, Germany

Received 30 November 1998/Returned for modification 22 March 1999/Accepted 9 April 1999

Seven complete genes and one incomplete gene for the biosynthesis of the glycopeptide antibiotic balhimycin were isolated from the producer, Amycolatopsis mediterranei DSM5908, by a reverse-cloning approach and characterized. Using oligonucleotides derived from glycosyltransferase sequences, a 900-bp glycosyltransferase gene fragment was amplified and used to identify a DNA fragment of 9,882 bp. Of the identified open reading frames, three (oxyA to -C) showed significant sequence similarities to cytochrome P450 monooxygenases and one (bhaA) showed similarities to halogenase, and the genes bgtfA to -C showed similarities to glycosyltransferases. Glycopeptide biosynthetic mutants were created by gene inactivation experiments eliminating oxygenase and glycosyltransferase functions. Inactivation of the oxygenase gene(s) resulted in a balhimycin mutant (SP1-1) which was not able to synthesize an antibiotically active compound. Structural analysis by high-performance liquid chromatography-mass spectrometry, fragmentation studies, and amino acid analysis demonstrated that these oxygenases are involved in the coupling of the aromatic side chains of the unusual heptapeptide. Mutant strain HD1, created by inactivation of the glycosyltransferase gene bgtfB, produced at least four different compounds which were not glycosylated but still antibiotically active.

* Corresponding author. Mailing address: Mikrobiologie/Biotechnologie, Eberhard-Karls-Universität Tübingen, Auf der Morgenstelle 28, D-72076 Tübingen, Germany. Phone: 49-7071-2976944. Fax: 49-7071-295979. E-mail: wowo{at}molbio.biol.biologie.uni-tuebingen.de.

dagger Present address: Universität des Saarlandes, Institut für Angewandte Mikrobiologie, Im Stadtwald, Gebäude 2, D-66041 Saarbrücken, Germany.

Antimicrobial Agents and Chemotherapy, July 1999, p. 1565-1573, Vol. 43, No. 7
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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