Lienhwalides: Unique Tropolone-Maleidride Hybrids from Hypoxylon lienhwacheense.

Abstract

Hypoxylon lienhwacheense, a fungal species with an unclear taxonomic placement within the Hypoxylaceae, presented a highly rare stromatal secondary metabolite profile. Isolation of its major stromatal constituents led to the discovery of a novel tropolone-maleidride hybrid molecule, lienhwalide A 5, in addition to the known cordyanhydride B 6, its new derivative 7, and binaphthalenetetraol 8. Unexpectedly, H. lienhwacheense produced in liquid cultures various lienhwalide A congeners 9-11. Their structures and relative configurations were elucidated using HRMS and NMR spectroscopy, with their absolute configurations determined using X-ray analysis of a semisynthetic brominated derivative of 9 and synthesising MTPA-esters of 11. Feeding experiments with 13C-labelled precursors (13C-methionine, 1-13C- and U-13C6-glucose) revealed insights into the biogenesis of tropolone and maleidride moieties, according to 13C couplings and INADEQUATE NMR data. Genome analysis identified two separate BGCs responsible for these moieties, and heterologous expression experiments provided further insights into the interplay of both clusters during the biosynthesis of these hybrid natural products. Remarkably, lienhwalides exhibited reduced toxicity and enhanced antibacterial selectivity compared to related fungal tropolones.