Generation of a cluster-free Streptomyces albus chassis strains for improved heterologous expression of secondary metabolite clusters

Abstract

Natural products are a rich source of potential drugs for many applications. Discovery of natural products through the activation of cryptic gene clusters encoding their biosynthetic pathways, engineering of those biosynthetic pathways and optimization of production yields often rely on the expression of these gene clusters in suitable heterologous host strains. Streptomyces albus J1074 provides high success rates of heterologous cluster expression with high levels of metabolite production, rapid growth and amenability to genetic manipulations. Here, we report the construction of S. albus chassis strains optimized for the discovery of natural products through heterologous expression of secondary metabolite clusters. 15 clusters encoding secondary metabolite biosynthetic pathways were deleted in the chromosome of S. albus Del14. This strain provides a substantially improved compound detection limit, owing to the lack of native secondary metabolites. Furthermore, the production yield of natural products heterologously expressed in S. albus Del14 was higher than in commonly used S. albus J1074 and S. coelicolor hosts. S. albus strains B2P1 and B4 were generated by introduction of additional phage phiC31 attB sites into the chromosome of S. albus Del14, allowing integration of up to four copies of a heterologous gene cluster. Amplification of gene clusters in the chromosome of the constructed strains further improved production yields of the encoded compounds. One cryptic cluster from Streptomyces spp. and two clusters from distantly related Frankia spp. strains were successfully activated in these new chassis strains, leading to the isolation of a new compound fralnimycin.

Keywords: Biosynthetic gene cluster; Chassis strain; Heterologous host; Natural products; Streptomyces.