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Bacteria of the genus  Saccharopolyspora  produce important polyketide antibiotics, including erythromycin A ( Sac. erythraea ) and spinosad ( Sac. spinosa ). We herein report the development of an industrial erythromycin-producing strain,  Sac .  erythraea  HOE107, into a host for the heterologous expression of polyketide biosynthetic gene clusters (BGCs) from other  Saccharopolyspora  species and related actinomycetes. To facilitate the integration of natural product BGCs and auxiliary genes beneficial for the production of natural products, the erythromycin polyketide synthase ( ery ) genes were replaced with two bacterial  attB  genomic integration sites associated with bacteriophages ϕC31 and ϕBT1. We also established a highly efficient conjugation protocol for the introduction of large bacterial artificial chromosome (BAC) clones into  Sac. erythraea  strains. Based on this optimized protocol, an arrayed BAC library was effectively transferred into  Sac. erythraea . The large spinosad gene cluster from  Sac. spinosa  and the actinorhodin gene cluster from  Streptomyces coelicolor  were successfully expressed in the  ery  deletion mutant. Deletion of the endogenous giant polyketide synthase genes  pkeA1 - pkeA4 , the product of which is not known, and the flaviolin gene cluster ( rpp ) from the bacterium increased the heterologous production of spinosad and actinorhodin. Furthermore, integration of pJTU6728 carrying additional beneficial genes dramatically improved the yield of actinorhodin in the engineered  Sac. erythraea  strains. Our study demonstrated that the engineered  Sac. erythraea  strains SLQ185, LJ161, and LJ162 are good hosts for the expression of heterologous antibiotics and should aid in expression-based genome-mining approaches for the discovery of new and cryptic antibiotics from  Streptomyces  and rare actinomycetes.

Engineering the Erythromycin-Producing Strain Saccharopolyspora erythraea HOE107 for the Heterologous Production of Polyketide Antibiotics