GliP, a multimodular nonribosomal peptide synthetase in Aspergillus fumigatus, makes the diketopiperazine scaffold of gliotoxin

The fungal metabolite gliotoxin has a redox-active disulfide bridge spanning carbons 3 and 6 of a diketopiperazine ( DKP) scaffold. The proposed DKP synthetase, GliP, from Aspergillus fumigatus Af293, is a three module ( A(1)-T-1-C-1-A(2)-T-2-C-2-T-3) 236 kDa protein that can be overproduced in soluble form in Escherichia coli. Once primed on its three thiolation domains with phosphopantetheine prosthetic groups, GliP activates and tethers L-Phe on T-1 and L-Ser on T-2, before generating the L-Phe-L-Ser-S-T-2 dipeptidyl enzyme intermediate. Release of the dipeptide as the cyclic DKP happens slowly both in wildtype GliP and in enzyme forms where C-2 and T-3 have been mutationally inactivated. The lack of a thioesterase domain in GliP may account both for the slow release and for the directed fate of intramolecular cyclization to create the DKP scaffold for subsequent elaboration to gliotoxin.