The natural product 'K-26'from an actinomycete is representative of an uninvestigated class of natural carbon-phosphorus bond containing compounds which incorporate a phosphonic acid analog of tyrosine. K- 26 possesses angiotensin converting enzyme (ACE) inhibitory activity with an IC50 value of 12.5 nM, comparable to the widely prescribed antihypertensive drug Captopril. K-26 is comprised of N-acetylated isoleucine, tyrosine, and the nonproteinogenic amino acid, (R)-1-amino-2-(4-hydroxyphenyl)ethylphosphonic acid (AHEP). Despite the potent hypotensive activity of K-26 and related compounds, the biosynthetic pathways by which K-26 and more specifically AHEP are biosynthesized still remain uncharacterized. The aromatic amino acid functionality of AHEP suggests that its origin lies in the shikimic acid pathway. However, it is difficult to rationalize how phosphoenolpyruvate mutase, the activity responsible for forming all previously studied phosphonate containing natural products, can be integrated into classical amino acid metabolism to generate AHEP. We describe preliminary data that demonstrate that AHEP is derived from tyrosine and that the C-P bond forming chemistry is unique. We hypothesize a mechanism for C-P bond formation and propose a series of studies to (1) define the basic building blocks and pathway of assembly of K-26, (2) identify the genes responsible for K-26 biosynthesis and (3) characterize the K-26 biosynthetic machinery, especially the C-P bond forming biochemistry. Impact on public health: Proposed research will be the first study of the biosynthesis of a unique class of compounds containing an aromatic phosphonic acid, a class which has been demonstrated to have potent metalloproteinase inhibitory activity. As such, this proposal will provide significant new tools for the discovery and recombinant biocatalytic generation of therapeutically useful novel phosphonate functional metalloproteinase inhibitors. Metalloproteinase inhibitors have potential to treat a large number of human illnesses including hypertension, cancer and osteoarthritis, among others.
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