Leinamycin (LNM), first discovered 30 years ago, exhibits unique biological activity and serves as a model system for the study of bacterial hybrid nonribosomal peptide synthetase (NRPS) – “AT-less” polyketide synthase (PKS) enzymology. Featuring a unique 1,3-dioxo-1,2-dithiolane moiety spiro-fused to an 18-membered macrolactam ring, LNM exhibits potent activity against tumors that are resistant to clinically important anticancer drugs. Upon reductive activation by cellular thiols, LNM exerts antitumor activity by an episulfonium ion-mediated DNA alkylation, a mode of action that is unprecedented among all DNA-damaging NPs. Furthermore, LNM is produced by an “AT-less” PKS assembly line and was the first PKS of this type to be characterized.
The LNM system has served as a fantastic model system for AT-less PKS and hybrid NRPS-PKS enzymology, and numerous chemical reactions have been characterized using LNM as a model system. Relevant examples that are general for other NPs include: (i) the trans-AT involved in loading substrates to each PKS module, (ii) beta-branching, and (iii) the enzymology surrounding the NRPS-PKS interface of hybrid NRPS-PKS systems.
Specifically for LNM, the chemistry involved in the incorporation of sulfur atoms into this scaffold is under intense investigation. A unique PKS didomain has been shown to be involved in sulfur incorporation, the DUF-SH (domain of unknown function – sulfhydralase). The genetic sequence of the DUF-SH didomain served as a basis for genome mining to discover new members of the LNM family of NPs (Figure 1A), which led to the discovery of the guangnanmycins (GNMs, Figure 1C). Using GNM as a model system, we have characterized the SH domain as a thiocysteine lyase, which implicates persulfide intermediates in LNM/GNM biosynthesis, and accounts for both of the sulfur atoms in the 1,3-dioxo-1,2-ditholane of LNM (Figure 1B), as well as the methyldisulfide moiety of GNM A (Figure 1C).
Using LNM and GNM as model systems for the LNM family of NPs, current efforts focus on: (i) characterizing the DUF-SH domains for sulfur incorporation into NP scaffolds by type I PKSs, (ii) mining actinomycetal genomes to discover new members of the LNM family of NPs, and (iii) unveiling the unprecedented chemistry and enzymology for sulfur-containing NP biosynthesis.
Figure 1. Overview of the diversity of the LNM family of NPs and the characterized sulfur chemistry. (A) Biosynthetic gene clusters from sequenced genomes within the Natural Product Discovery Center demonstrating the genetic and chemical diversity available. (B) Structure of LNM. (C) Thiocysteine lyase activity of the SH domain from the PKS assembly line responsible for the biosynthesis of GNM A, leading to the realization that the SH domains function as thiocysteine lyases.