Toward Improvement of Erythromycin A Production in an IndustrialSaccharopolyspora erythraea Strain via Facilitation of Genetic Manipulation with an Artificial attB Site for Specific Recombination
Open Access
- 1 November 2011
- journal article
- Published by American Society for Microbiology in Applied and Environmental Microbiology
- Vol. 77 (21), 7508-7516
- https://doi.org/10.1128/aem.06034-11
Abstract
Large-scale production of erythromycin A (Er-A) relies on the organism Saccharopolyspora erythraea , in which lack of a typical attB site largely impedes the application of phage ΦC31 integrase-mediated recombination into site-specific engineering. We herein report construction of an artificial attB site in an industrial S. erythraea strain, HL3168 E3, in an effort to break the bottleneck previously encountered during genetic manipulation mainly from homologous or unpredictable nonspecific integration. Replacement of a cryptic gene, nrps1-1 , with a cassette containing eight attB DNA sequences did not affect the high Er-producing ability, setting the stage for precisely engineering the industrial Er-producing strain for foreign DNA introduction with a reliable conjugation frequency. Transfer of either exogenous or endogenous genes of importance to Er-A biosynthesis, including the S -adenosylmethionine synthetase gene for positive regulation, vhb for increasing the oxygen supply, and two tailoring genes, eryK and eryG , for optimizing the biotransformation at the late stage, was achieved by taking advantage of this facility, allowing systematic improvement of Er-A production as well as elimination of the by-products Er-B and Er-C in fermentation. The strategy developed here can generally be applicable to other strains that lack the attB site.Keywords
This publication has 37 references indexed in Scilit:
- Streptomyces and Saccharopolyspora hosts for heterologous expression of secondary metabolite gene clustersJournal of Industrial Microbiology & Biotechnology, 2010
- Natural products to drugs: natural product-derived compounds in clinical trialsNatural Product Reports, 2008
- Genetic Modulation of the Overexpression of Tailoring Genes eryK and eryG Leading to the Improvement of Erythromycin A Purity and Production in Saccharopolyspora erythraea FermentationApplied and Environmental Microbiology, 2008
- Development of an enhanced separation of erythromycin and its related substances by liquid chromatographyJournal of Pharmaceutical and Biomedical Analysis, 2007
- Molecular engineering approaches to peptide, polyketide and other antibioticsNature Biotechnology, 2006
- Directing BiosynthesisScience, 2006
- Genetic improvement of processes yielding microbial productsFEMS Microbiology Reviews, 2006
- The Streptomyces Genome Contains Multiple Pseudo- attB Sites for the φC31-Encoded Site-Specific Recombination SystemJournal of Bacteriology, 2002
- Genetic engineering of an industrial strain of Saccharopolyspora erythraea for stable expression of the Vitreoscilla haemoglobin gene (vhb)Microbiology, 1998
- Plasmid cloning vectors for the conjugal transfer of DNA from Escherichia coli to Streptomyces spp.Gene, 1992