The Fumitremorgin Gene Cluster of Aspergillus fumigatus: Identification of a Gene Encoding Brevianamide F Synthetase

Abstract

A gene encoding a putative dimodular nonribosomal peptide synthetase (NRPS) was identified within a gene cluster of Aspergillus fumigatus, a species reported to produce fumitremorgins and other prenylated alkaloids. The gene was deleted and overexpressed in the genome reference strain Af293, and was also expressed in the naïve host Aspergillus nidulans, which lacks the equivalent gene cluster. While neither fumitremorgins nor the dipeptide brevianamide F (cyclo-L-Trp-L-Pro), an early intermediate, were detected in wild-type and deletion strains of A. fumigatus, brevianamide F accumulated in fungal cultures following increased expression of the NRPS gene in both A. fumigatus and A. nidulans. We conclude that the gene Afu8g00170, named ftmA, encodes the NRPS brevianamide synthetase. Brevianamide F is the precursor of a variety of fungal prenylated alkaloids with biological activity, including fumitremorgins A, B and C and tryprostatin B.

CdpC2PT, a reverse prenyltransferase from Neosartorya fischeri with a distinct substrate preference from known C2-prenyltransferases

“…CdpC2PT could be responsible for the two prenylation steps. Formation of the diketopiperazine ring could be catalysed by a nonribosomal peptide synthetase (Ames & Walsh, 2010;Maiya et al, 2006;Yin et al, 2009b). Inspection of the genome sequence of N. fischeri revealed the presence of a bimodular non-ribosomal peptide synthetase gene NFIA_043670 close to NFIA_043650 (Fig.…”

Genome mining and biosynthesis of fumitremorgin-type alkaloids in ascomycetes

“…The fumitremorgin/verruculogen cluster was very likely not expressed in A. fumigatus Af293; therefore, no secondary metabolites of this cluster could be detected in this strain. 33 In contrast, a number of intermediates including tryprostatins A (3) and B (2), fumitremorgin C (8), 12,13-dihydroxyfumitremorgin C, fumitremorgin B (9) as well as verruculogen (11) were isolated from A. fumigatus BM939. 5 No data on the expression of the gene clusters in A. fumigatus A1163 and N. fischeri NRRL181 are available.…”

“…Functional proof of the biosynthetic genes and gene cluster of fumitremorgin-type alkaloids was carried out by gene inactivation experiments in the producing strain BM939, 31 by identification of the accumulated secondary metabolites after heterologous expression in Aspergillus strains 33,34 or by biochemical investigation of the recombinant enzymes from Escherichia coli or Saccharomyces cerevisiae. 27,31,32,35 In total, functions of seven genes have been proven experimentally.…”

“…On the basis of the obtained bioinformatic, molecular biological and biochemical results, the following biosynthetic pathway has been postulated ( Figure 3). 1 The biosynthesis of fumitremorgin-type alkaloids begins by condensation of the two amino acids L-tryptophan and L-proline to brevianamide F, catalyzed by the non-ribosomal peptide synthetase FtmPS (also termed FtmA), which was proven by heterologous overexpression of the NRPS gene ftmPS in Aspergillus and identification of the accumulated product brevianamide F. 33 Brevianamide F is then prenylated by the prenyltransferase FtmPT1/FtmB in the presence of dimethylallyl diphosphate, resulting in the formation of tryprostatin B, as demonstrated by using the recombinant FtmPT1. 27 By gene inactivation in A. fumigatus BM939 and heterologous expression in S. cerevisiae, Kato et al 31 showed that the three cytochrome P450 enzymes, FtmP450-1/FtmC, FtmP450-2/FtmE and FtmP450-3/FtmG, are responsible for the conversion of tryprostatin B (2) to 6-hydroxytryprostatin B, tryprostatin A (3) to fumitremorgin C (8) and fumitremorgin C (8) to 12,13-dihydroxyfumitremorgin C, respectively.…”

The first filamentous fungal genome sequences: Aspergillus leads the way for essential everyday resources or dusty museum specimens?

“…The focused research within secondary metabolites, such as statin biosynthesis in A. terreus, provides expertise and resources with a reason to address specific pathways and classes of genes. The role of toxins in A. fumigatus pathogenicity is another longstanding research question and the genome sequence has already aided location of biosynthetic genes for some of these toxins (Maiya et al, 2006).…”