Computational identification and analysis of orphan assembly-line polyketide synthases

Abstract

The increasing availability of DNA sequence data offers an opportunity for identifying new assembly-line polyketide synthases (PKSs) that produce biologically active natural products. We developed an automated method to extract and consolidate all multimodular PKS sequences (including hybrid PKS/non-ribosomal peptide synthetases) in the National Center for Biotechnology Information (NCBI) database, generating a non-redundant catalog of 885 distinct assembly-line PKSs, the majority of which were orphans associated with no known polyketide product. Two in silico experiments highlight the value of this search method and resulting catalog. First, we identified an orphan that could be engineered to produce an analog of albocycline, an interesting antibiotic whose gene cluster has not yet been sequenced. Second, we identified and analyzed a hitherto overlooked family of metazoan multimodular PKSs, including one from Caenorhabditis elegans. We also developed a comparative analysis method that identified sequence relationships among known and orphan PKSs. As expected, PKS sequences clustered according to structural similarities between their polyketide products. The utility of this method was illustrated by highlighting an interesting orphan from the genus Burkholderia that has no close relatives. Our search method and catalog provide a community resource for the discovery of new families of assembly-line PKSs and their antibiotic products.

Figure 1

Summary of workflow. A full color version of this figure is available at The Journal of Antibiotics journal online.

Figure 2

Sequence similarity relationships among PKSs involved in the biosynthesis of known polyketide natural products. The amino-acid sequences of 62 representative assembly-line PKSs and PKS–NRPSs, plus the three orphan PKSs highlighted in this report, were compared in a pairwise manner using a gene-cluster similarity score. Scale bar displays the distance between gene clusters (between 0 and 1). The label for each PKS lists the Genbank ID and cluster number, date the sequence was deposited in NCBI, number of KS, AT, C and A domains, and sequence origin as listed in NCBI. The three orphans highlighted in the text are displayed in red. A full color version of this figure is available at The Journal of Antibiotics journal online.

Figure 3

Cumulative number of non-redundant PKS gene clusters in NCBI over time: data were collected through the first half of 2013. A full color version of this figure is available at The Journal of Antibiotics journal online.

Figure 4

An orphan cluster that may produce a natural product that is structurally analogous to albocycline, a potent antibiotic derived from Streptomyces sp. 6–31, whose encoding gene cluster is unknown. This cluster could be engineered to produce albocycline analogs. A full color version of this figure is available at The Journal of Antibiotics journal online.

Figure 5

Predicted PKS assembly line for a cluster from Burkholderia spp. that does not appear related to any known biosynthetic clusters A full color version of this figure is available at The Journal of Antibiotics journal online.

Figure 6

(a) Predicted structure of a hybrid PKS/NRPS assembly-line gene in C. elegans. (b) Clade of nematode homologs of the hybrid PKS/NRPS C. elegans gene in part A, clustered according to our heuristic PKS sequence similarity score (see also Supplementary Figure 5). A full color version of this figure is available at The Journal of Antibiotics journal online.