Unlocking New Prenylation Modes: Azaindoles as a New Substrate Class for Indole Prenyltransferases

Aza-substitution, the replacement of aromatic CH groups with nitrogen atoms, is an established medicinal chemistry strategy for increasing solubility, but current methods of accessing functionalized azaindoles are limited. In this work, indole-alkylating aromatic prenyltransferases (PTs) were explored as a strategy to directly functionalize azaindole-substituted analogs of natural products. For this, a series of aza-L-tryptophans (Aza-Trp) featuring N-substitution of every aromatic CH position of the indole ring and their corresponding cyclic Aza-L-Trp-L-proline dipeptides (Aza-CyWP), were synthesized as substrate mimetics for the indole-alkylating PTs FgaPT2, CdpNPT, and FtmPT1. We then demonstrated most of these substrate analogs were accepted by a PT, and the regioselectivity of each prenylation was heavily influenced by the position of the N-substitution. Remarkably, FgaPT2 was found to produce cationic N-prenylpyridinium products, representing not only a new substrate class for indole PTs but also a previously unobserved prenylation mode. The discovery that nitrogenous indole bioisosteres can be accepted by PTs thus provides access to previously unavailable chemical space in the search for bioactive indolediketopiperazine analogs. Enzyme catalysis: Five isomers of aza-tryptophan and the corresponding series of cyclic Aza-tryptophan-proline dipeptides were tested as unnatural substrates for three well-known indole-prenylating enzymes, FgaPT2, CdpNPT, and FtmPT1. Most substrates were found to produce prenylated products, which revealed isomer-dependent regioselectivity and a previously unreported class of cationic N-prenyl pyridinium products.image