title: Toward improvement of erythromycin A production in an industrial Saccharopolyspora erythraea strain via facilitation of genetic manipulation with an artificial attB site for specific recombination.
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.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Jiequn
authors.initials: J
authors.lastname: Wu
authors.forename: Qinglin
authors.initials: Q
authors.lastname: Zhang
authors.forename: Wei
authors.initials: W
authors.lastname: Deng
authors.forename: Jiangchao
authors.initials: J
authors.lastname: Qian
authors.forename: Siliang
authors.initials: S
authors.lastname: Zhang
authors.forename: Wen
authors.initials: W
authors.lastname: Liu
mesh_terms: D001287:Attachment Sites, Microbiological; D003227:Conjugation, Genetic; D004917:Erythromycin / Q000378:metabolism*; D005818:Genetic Engineering / Q000379:methods*; D005827:Genetics, Microbial / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D030781:Organisms, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D016930:Saccharopolyspora / Q000235:genetics / Q000378:metabolism*
chemical_list: D004917:Erythromycin