Echinocandin B biosynthesis: a biosynthetic cluster from Aspergillus nidulans NRRL 8112 and reassembly of the subclusters Ecd and Hty from Aspergillus pachycristatus NRRL 11440 reveals a single…

Wolfgang Hüttel, Loubna Youssar, Björn A. Grüning, Stefan Günther, Katharina G. Hugentobler

Echinocandins are nonribosomal lipopeptides produced by ascommycete fungi. Due to their strong inhibitory effect on fungal cell wall biosynthesis and lack of human toxicity, they have been developed to an important class of antifungal drugs. Since 2012, the biosynthetic gene clusters of most of the main echinocandin variants have been characterized. Especially the comparison of the clusters allows a deeper insight for the biosynthesis of these complex structures. In the genome of the echinocandin B producer Aspergillus nidulans NRRL 8112 we have identified a gene cluster (Ani) that encodes echinocandin biosynthesis. Sequence analyses showed that Ani is clearly delimited from the genomic context and forms a monophyletic lineage with the other echinocandin gene clusters. Importantly, we found that the disjunct genomic location of the echinocandin B gene cluster in A. pachycristatus NRRL 11440 on two separate subclusters, Ecd and Hty, at two loci was likely an artifact of genome misassembly in the absence of a reference sequence. We show that both sequences can be aligned resulting a single cluster with a gene arrangement collinear compared to other clusters of Aspergillus section Nidulantes. The reassembled gene cluster (Ecd/Hty) is identical to a putative gene cluster (AE) that was previously deposited at the NCBI as a sequence from A. delacroxii NRRL 3860. PCR amplification of a part of the gene cluster resulted a sequence that was very similar (97 % identity), but not identical to that of AE. The Echinocandin B biosynthetic cluster from A. nidulans NRRL 8112 (Ani) is particularly similar to that of A. pachycristatus NRRL 11440 (Ecd/Hty). Ecd/Hty was originally reported as two disjunct sub-clusters Ecd and Hty, but is in fact a continuous sequence with the same gene order as in Ani. According to sequences of PCR products amplified from genomic DNA, the echinocandin B producer A. delacroxii NRRL 3860 is closely related to A. pachycristatus NRRL 11440. A PCR-product from the gene cluster was very similar, but clearly distinct from the sequence published for A. delacroxii NRRL 3860 at the NCBI (No. AB720074). As the NCBI entry is virtually identical with the re-assembled Ecd/Hty cluster, it is likely that it originates from A. pachycristatus NRRL 11440 rather than A. delacroxii NRRL 3860.