### **The Natural Product Domain Seeker version 2 (NaPDoS2): Relating ketosynthase phylogeny to biosynthetic function** ---------- *Background:* $\space$ The Natural Product Domain Seeker (NaPDoS) web tool detects and classifies ketosynthase (KS) and condensation (C) domains using a phylogeny-based functional classification scheme to make predictions about the diversity of polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) gene cluster diversity. The use of domain sequence signature tags makes NaPDoS particularly useful for analyzing incomplete and/or non-contiguous biosynthetic gene clusters from poorly assembled genomes and metagenomes, as well as intron-punctuated eukaryotic genomes. Here we introduce version 2 of the webtool (NaPDoS2), which is publicly available at http://napdos.ucsd.edu/napdos2. Taxonomic and functional coverage has been greatly expanded in NaPDoS2 to incorporate 1417 new KS sequences representing 41 KS classes and subclasses, including new subclass assignments for type II PKSs, along with increased representation of eukaryotic KSs. User interface improvements and workflow modifications greatly accelerate web tool run times, allowing much larger data sets to be analyzed. Statistical validation tests have been performed to evaluate program performance, establishing default parameters that maximize KS detection and classification accuracy while minimizing false positives on a variety of input data types. The rapid insights NaPDoS2 provides into PKS biosynthetic potential are demonstrated by extensive application use cases drawn from genome, metagenome, and PCR amplicon datasets. These examples show how this tool can be used to guide the discovery of gene clusters involved in the biosynthesis of compounds within specific structure classes and aid in identifying divergent KS or C domains that may be associated with new biosynthetic mechanisms. ---------- *Reference:* $\space$ Leesa J Klau*, Sheila Podell*, Kaitlin E Creamer*, Alyssa M Demko, Hans W Singh, Eric E. Allen, Bradley S Moore, Nadine Ziemert, Anne Catrin Letzel, Paul R Jensen. **The Natural Product Domain Seeker version 2 (NaPDoS2) webtool relates ketosynthase phylogeny to biosynthetic function**. (2022). Journal of Biological Chemistry https://doi.org/10.1016/j.jbc.2022.102480 ---------- ![NaPDoS2 graphical abstract][1] ---------- ### **Phylogenetic analyses**: Sequence files in `.fasta` format, alignment files in `.fasta` format, and tree files in `.nhx` format generated from phylogenetic analyses corresponding to **Figures 1-3** and **Supplemental Figures S5-S7**. --- - **Figure 1, Figure S7:** Maximum likelihood phylogeny of 414 KS sequences and three outgroup sequences. Sequences aligned using MAFFT and trimmed using TrimA1. Tree constructed using FastTree with 1000 bootstraps, supports estimated using Booster. Sequences: `1-01-KSdb13_refseqs_thiolaseOG_270714.fasta` Alignment: `1-02-RT-OG-MAFFT-trimA1.fasta` Tree: `1-05-RT-OG-MAFFT-trimA1_Booster__Tree_with_normalized_supports__tbe_norm_tree.nhx` --- - **Figure 3, Figure S8:** Maximum likelihood phylogeny of 212 type II KS sequences and three outgroup sequences. Sequences aligned using MAFFT and trimmed using TrimA1. Tree constructed using FastTree with 1000 bootstraps, supports estimated using Booster. Sequences: `2-01-KSdb13_typeIIall_outgroup_FASs_210830.fasta` Alignment: `2-05-T2KStree-OG-MAFFT-trimA1_trimAl_Output_Fasta.fasta` Tree: `2-09-T2KStree-OG-MAFFT-trimA1_Booster__Tree_with_normalized_supports__tbe_norm_tree.nhx` --- - **Figure 4, Figure S9:** Maximum likelihood phylogeny of concatenated KS alpha and beta subunits from 59 type II aromatic PKS. Sequences aligned using MAFFT and trimmed using TrimA1. Tree constructed using FastTree with 1000 bootstraps, supports estimated using Booster. Sequences: `3-01-KSdb13_typeIIaromatic_concat_210830.fasta` Alignment: `3-05-T2aroKStree-concat-MAFFT-trimA1_trimAl_Output_Fasta.fasta` Tree: `3-09-T2aroKStree-concat-MAFFT-trimA1_Booster__Tree_with_normalized_supports__tbe_norm_tree.nhx` --- [1]: https://files.osf.io/v1/resources/uzhcp/providers/osfstorage/61f3966079e27202cb20e222?mode=render