Nonribosomal peptide synthetases (NRPSs) are multifunctional enzymes that produce a wide array of bioactive peptides. Here we show that a single tryptophan-to-serine mutation in phenylalanine-specific NRPS adenylation domains enables the efficient activation of non-natural aromatic amino acids functionalized with azide and alkyne groups. The resulting 10(5)-fold switch in substrate specificity was achieved without appreciable loss of catalytic efficiency. Moreover, the effective communication of the modified A domains with downstream modules in dipeptide synthetases permitted incorporation of O-propargyl-L-tyrosine into diketopiperazines both in vitro and in vivo, even in the presence of competing phenylalanine. Because azides and alkynes readily undergo bioorthogonal click reactions, reprogramming NRPSs to accept non-natural amino acids that contain these groups provides a potentially powerful means of isolating, labeling, and modifying biologically active peptides.

N-Heterocyclic carbene catalyzed aza-Claisen annulations of enals or their α'-hydroxyenone surrogates with vinylogous amides afford dihydropyridinones. The reaction proceeds with a broad range of substrates, and no nitrogen protecting group is required.

Nonribosomal peptide synthetases (NRPSs) are multifunctional enzymes that produce a wide array of bioactive peptides. Here we show that a single tryptophan-to-serine mutation in phenylalanine-specific NRPS adenylation domains enables the efficient activation of non-natural aromatic amino acids functionalized with azide and alkyne groups. The resulting 10 5 -fold switch in substrate specificity was achieved without appreciable loss of catalytic efficiency. Moreover, the effective communication of the modified A domains with downstream modules in dipeptide synthetases permitted incorporation of O-propargyl-l-tyrosine into diketopiperazines both in vitro and in vivo, even in the presence of competing phenylalanine. Because azides and alkynes readily undergo bioorthogonal click reactions, reprogramming NRPSs to accept non-natural amino acids that contain these groups provides a potentially powerful means of isolating, labeling, and modifying biologically active peptides.[**] We thank Prof. Mohamed A. Marahiel (Philipps Universität Marburg) for supplying strain HM0079 and plasmids pSU18_tycA and pTrc99a_tycB1, and Prof. Hans-Martin Fischer (ETH Zürich) for assistance with radiochemical experiments. We are also grateful to Prof. Chaitan Khosla (Stanford University) for valuable discussions.

The catalytic kinetic resolution of cyclic amines with achiral N-heterocyclic carbenes (NHC) and chiral hydroxamic acids has emerged as a promising method to obtain enantioenriched amines with high selectivity factors. In this report, we describe the catalytic kinetic resolution of disubstituted piperdines with practical selectivity factors (s up to 52) in which we uncovered an unexpected and pronounced conformational effect resulting in disparate reactivity and selectivity between the cis and trans-substituted piperidine isomers. Detailed experimental and computational (DFT) studies of the kinetic resolution of various disubstituted piperidines revealed a strong preference for the acylation of conformers in which the α-substituent occupies the axial position. This work provides further experimental and computational support for the concerted 7-member transition state model for acyl transfer reagents and expands the scope and functional group tolerance of the secondary amine kinetic resolution.

The current laboratory practices of organic synthesis are labor intensive, impose safety and environmental hazards, and hamper the implementation of artificial intelligence guided drug discovery. Using a combination of reagent...