Computational protocol: Experimental and Metabolic Modeling Evidence for a Folate-Cleaving Side-Activity of Ketopantoate Hydroxymethyltransferase (PanB)

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Protocol publication

[…] The BLAST tools (Altschul et al., ) and resources at NCBI (http://www.ncbi.nlm.nih.gov/) were routinely used. Sequences were aligned using Clustal Omega (Li et al., ) or Multalin (Corpet, ). Sequence logos were generated using WebLogo3 (http://weblogo.threeplusone.com/) (Crooks et al., ) or with the logo comparison tool at http://www.twosamplelogo.org/ (Vacic et al., ). Phylogenetic distribution was analyzed in the SEED database (Overbeek et al., ). Results are available in the “Folate_biosynthesis” subsystem on the SEED server (http://pubseed.theseed.org/SubsysEditor.cgi?page=ShowSubsystem&subsystem=Folate_Biosynthesis). Physical clustering was analyzed with the SEED subsystem coloring tool or the SeedViewer Compare Regions tool (Overbeek et al., ). A subset of FolK sequences from ~1000 representative genomes was extracted from SEED via a perl API query (Disz et al., ). An alignment was done using Muscle (http://www.drive5.com/muscle/) (Edgar, ) on the command line with default settings. The trees were made with MEGA6 (Tamura et al., ) using Maximum Likelihood, Neighbor Joining, and UPGMA methods. The trees were visualized with the ETE2 python package (Huerta-Cepas et al., ). The physical clustering information was added to the tree after writing a python code that extracted if folK was within 10 coding sequences of the folB gene, the panB gene or both in a given genome. PDB (www.rcsb.org) (Berman et al., ) was used to visualize structures and ligand binding sites. […]

Pipeline specifications

Software tools Clustal Omega, MultAlin, WebLogo, MUSCLE, MEGA
Applications Phylogenetics, Nucleotide sequence alignment, Genome data visualization
Organisms Escherichia coli
Chemicals Carbon, Folic Acid