FK506 is a secondary metabolite with a potent immunosuppressant activity, currently registered for use after organ transplantation. Nevertheless, many aspects of its biosynthesis remained unclear. In this article, we report the identification of a previously unknown region of the FK506 gene cluster from S. tsukubaensis NRRL 18488 containing genes encoding provision of the unusual building blocks. We identified a group of genes, termed the “all” subcluster encoding biosynthesis of the extender unit allylmalonyl-CoA which provides the allyl group at C21 carbon atom of FK506. Interestingly, a small independent enzyme diketide synthase was identified to be involved in the biosynthesis of the allylmalonyl-CoA extender unit. Based on the newly identified genes researchers of Acies Bio and Lek Sandoz prepared a patent application to be filed at EPO, as a part of the work described in this research article is directly applicable for FK506 process improvement.
COBISS.SI-ID: 3754104
In this article we report the development of a novel bioprocess which enables exclusive biosynthesis/production of the immunosuppressant FK506 (tacrolimus) without simultaneous production of two structurally very similar compounds FK520 and dihydro-FK506. Traditional bioprocesses for production of FK506 result in the levels of these two by-products between 10 and 20% of the produced FK506, which due to great structural similarity of the compounds hampers the isolation of FK506 for use in pharmaceutical industry. Purification using industrial HPLC greatly increases the price of the final product as well as the environmental burden due to large amounts of organic solvents used. The novel process is based on the targeted genetic modification of the strain Streptomyces tsukubaensis NRRL18488 in which the supply of building blocks for polyketide synthase was abolished. Fermentation broths of the modified strain were then supplemented with a synthetic analogue of the allylmalonyl-CoA extender unit. Based on these results, Sandoz/Lek d.d. filed an application at the European Patent Office (EP2272963).
COBISS.SI-ID: 3988600
In this article was developed a robust and versatile reporter system based on the rppA gene from Saccharopolyspora erythraea. The applicability of the reporter system has been demonstrated by expressing the rppA gene under the control of the heterologous promoters actII-ORF4/PactI, ermE and its upregulated variant ermE*. The simplicity and robustness of the system, demonstrated even in industrial settings, shows great potential for wider use in different microbial hosts and applications, and may thus represent a new generic and versatile tool useful to a wider scientific community.
COBISS.SI-ID: 3811960