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Applied and Environmental Microbiology, April 2005, p. 1829-1835, Vol. 71, No. 4
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.4.1829-1835.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Rapid Engineering of the Geldanamycin Biosynthesis Pathway by Red/ET Recombination and Gene Complementation

Leandro Vetcher, Zong-Qiang Tian, Robert McDaniel, Andreas Rascher, W. Peter Revill, C. Richard Hutchinson, and Zhihao Hu*

Kosan Biosciences Inc., Hayward, California

Received 18 August 2004/ Accepted 5 November 2004

Genetic manipulation of antibiotic producers, such as Streptomyces species, is a rational approach to improve the properties of biologically active molecules. However, this can be a slow and sometimes problematic process. Red/ET recombination in an Escherichia coli host has permitted rapid and more versatile engineering of geldanamycin biosynthetic genes in a complementation plasmid, which can then be readily transferred into the Streptomyces host from which the corresponding wild type gene(s) has been removed. With this rapid Red/ET recombination and gene complementation approach, efficient gene disruptions and gene replacements in the geldanamycin biosynthetic gene cluster have been successfully achieved. As an example, we describe here the creation of a ketoreductase 6 null mutation in an E. coli high-copy-number plasmid carrying gdmA2A3 from Streptomyces hygroscopicus NRRL3602 and the subsequent complementation of a gdmA2A3 deletion host with this plasmid to generate a novel geldanamycin analog.

* Corresponding author. Mailing address: Kosan Biosciences Inc., 3832 Bay Center Place, Hayward, CA 94545. Phone: (510) 731-5311. Fax: (510)732-8401. E-mail: hu{at}kosan.com.

Applied and Environmental Microbiology, April 2005, p. 1829-1835, Vol. 71, No. 4
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.4.1829-1835.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Perlova, O., Fu, J., Kuhlmann, S., Krug, D., Stewart, A. F., Zhang, Y., Muller, R. (2006). Reconstitution of the Myxothiazol Biosynthetic Gene Cluster by Red/ET Recombination and Heterologous Expression in Myxococcus xanthus. Appl. Environ. Microbiol. 72: 7485-7494 [Abstract] [Full Text]  


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