Genome Mining of the Biocontrol Fungus Trichoderma afroharzianum T22 Led to the Discovery of Dehydrobutyrine-Containing Lipopeptides

Abstract

Elucidating natural products encoded by cryptic biosynthetic gene clusters (BGCs) in agriculturally important biocontrol fungi such as Trichoderma afroharzianum T22 (ThT22) could lead to the discovery of new compounds that may be explored as potential agrochemicals. Herein, we discovered new lipopeptide natural products, trichopeptide A–D, via genome mining and heterologous reconstitution of the tep BGC from ThT22. Comprehensive NMR, high-resolution mass spectrometry (HRMS), tandem mass spectrometry (MS/MS), Marfey’s analysis, and chemical synthesis experiments established the structures of trichopeptides as linear pentapeptides acylated with a conjugated triene acid at the N-terminus. A notable structural feature of trichopeptides is the presence of a nonproteinogenic amino acid residue, (Z)-dehydrobutyrine, in the linear peptide. The residue is proposed to be derived from threonine by the action of a noncanonical dehydrating condensation domain (DhC) in the nonribosomal peptide synthetase (NRPS) TepB. This represents only the second example in which a fungal NRPS is responsible for the biosynthesis of dehydroamino acids in a peptide product. Our results expand the chemical inventory of an agriculturally important fungus and pave the way for the future investigation of possible plant–ThT22 interactions mediated by lipopeptides.