Fig. 4

Calpinactam production in M. alpina and transgenic A. niger ∆fwnA mutants tJMW06 and tLK07. A Chemical structure and calculated MS/MS fragments of calpinactam. B Schematic representation of the integration event of the five calA fragments into the genomic fwnA locus of A. niger tLK01. The pearls on a string represent the domain structure of the 6-module NRPS CalA and include: C, condensation domain; A, adenylation domain; T, thiolation domain; E/C dual epimerization/condensation domain; TE, thioesterase. Note that the 1st (starter) C domain is truncated and inactive. C Extracted ion chromatograms of the synthesized calpinactam standard and metabolic extracts from various fungal mycelia. M. alpina ATCC32222 was cultivated on MEP, whilst transgenic A. niger tJMW06 (calA expressing) and tLK07 (empty vector control) was cultivated in Aspergillus Minimal Medium (AMM) under inducing (+ doxycycline) or non-inducing (− doxycycline) conditions. MS data was monitored in positive ionization mode at m/z 768.6844 [M + H]⁺. D MS/MS spectra highlighting the specific daughter ion fragments of calpinactam of the authentic standard and the metabolic extracts from M. alpina ATCC32222 and transgenic A. niger tJMW06