These are preliminary reports that have not been peer-reviewed. They should not be regarded as conclusive, guide clinical practice/health-related behavior, or be reported in news media as established information. For more information, please see our FAQs.
Preprints are manuscripts made publicly available before they have been submitted for formal peer review and publication. They might contain new research findings or data. Preprints can be a draft or final version of an author's research but must not have been accepted for publication at the time of submission.
submitted on 31.03.2020 and posted on 01.04.2020by Mengbin Chen, Chun-Ting Liu, Yi Tang
Pyridoxal phosphate (PLP)-dependent enzymes can catalyze various transformations of amino acids at alpha, beta, and gamma positions. These versatile enzymes are prominently involved in the biosynthesis of nonproteinogenic amino acids as building blocks of natural products, and are attractive biocatalysts. Here, we report the discovery of a two-step enzymatic synthesis of (2S, 6S)-6-methyl pipecolate 1, from the biosynthetic pathway of indole alkaloid citrinadin. The key enzyme CndF is PLP-dependent and catalyzes synthesis of (S)-2-amino-6-oxoheptanoate 3 that is in equilibrium with the cyclic Schiff base. The second enzyme CndE is a stereoselective imine reductase that gives 1. Biochemical characterization of CndF showed this enzyme performs gamma-elimination of O-acetyl L-homoserine to generate the vinylglycine ketimine, which is subjected to nucleophilic attack by acetoacetate to form the new Cgamma-Cdelta bond in 3 and complete the gamma-substitution reaction. CndF displays substrate promiscuity towards different beta-keto carboxylate and esters. Using a recombinant Aspergillus strain expressing CndF and CndE, feeding various alkyl-beta-keto esters led to the biosynthesis of 6-substituted L-pipecolates. The discovery of CndF expands the repertoire of reactions that can be catalyzed by PLP-dependent enzymes.