Cloning, nucleotide sequence, and heterologous expression of the biosynthetic gene cluster for R1128, a non-steroidal estrogen receptor antagonist - Insights into an unusual priming mechanism

R1128 substances are anthraquinone natural products that were previously re ported as non-steroidal estrogen receptor antagonists with in vitro and in vivo potency approaching that of tamoxifen, From a biosynthetic viewpoint, these polyketides possess structurally interesting features such as an unus ual primer unit that are absent in the well studied anthracyclic and tetrac yclic natural products. The entire R1128 gene cluster was cloned and expres sed in Streptomyces lividans, a genetically well developed heterologous hos t. In addition to R1128C, a novel optically active natural product, designa ted HU235, was isolated. Nucleotide sequence analysis of the biosynthetic g ene cluster revealed genes encoding two ketosynthases, a chain length facto r, an acyl transferase, three acetyl-CoA carboxylase subunits, two cyclases , two oxygenases, an amidase, and remarkably, two acyl carrier proteins. Fe eding studies indicate that the unusual 4-methylvaleryl side chain of R1128 C is derived from valine, Together with the absence of a dedicated ketoredu ctase, dehydratase, or enoylreductase within the R1128 gene cluster, this s uggests a functional link between fatty acid biosynthesis and R1128 biosynt hesis in the engineered host. Specifically, we propose that the R1128 synth ase recruits four subunits from the endogenous fatty acid synthase during t he biosynthesis of this family of pharmacologically significant natural pro ducts.