TioS T-TE--a prototypical thioesterase responsible for cyclodimerization of the quinoline- and quinoxaline-type class...

The family of chromodepsipeptides constitutes a class of structurally related pseudosymmetrical peptidolactones and peptidothiolactones synthesized by nonribosomal peptide synthetases. The chromodepsipeptides, which are analogous to the extensively characterized echinomycin, attain their DNA-bisintercalating properties from chromophore moieties attached to the N-termini of the oligopeptide chain. Thiocoraline, a quinoline-substituted DNA-bisintercalator isolated from marine actinomycetes, is a two-fold symmetric octathiodepsipeptide currently undergoing preclinical trials phase II. In the present study, the excised peptide cyclase TioS T-TE (thiolation-thioesterase bidomain) was employed as a general catalyst for the in vitro generation of thiocoraline analogs. TioS T-TE is capable of catalyzing ligation and the subsequent cyclization of tetrapeptidyl-thioester substrates, circumventing the demanding synthesis of octapeptidyl substrates. The general importance of several amino acid residues within the tetrapeptide was evaluated and revealed new insights with respect to the iterative mechanism utilized by the thioesterase. Additionally, substrate tolerance towards the cyclizing nucleophile allows the formation of macrolactones instead of the native macrothiolactones. Several thiocoraline analogs were isolated and investigated for DNA-bisintercalation activity. Relaxed substrate specificity regarding the chromophore moiety enables the chemoenzymatic synthesis of the quinoxaline- and quinoline-type class of chromodepsipeptides. TioS T-TE is the first nonribosomal peptide synthetase-derived thioesterase, capable of macrothiolactonization and macrolactonization, working in an iterative manner.