Fig 1.
novoStoic2.0: overall workflow for the integrated pathway design, including tools for optimal overall stoichiometry of conversion, optStoic.
De novo pathway design using optimal stoichiometry, novoStoic. Thermodynamic assessment of reaction steps using dGPredictor and rank-ordering of known enzymes for novel substrate activity to identify starting enzymes for novel steps using EnzRank.
Fig 2.
Annotated workflow of the home page of the novoStoic2.0 user interface.
This page provides an option for users to select specific tasks, standalone as well as in the given workflow, to include all the tools for designing pathways.
Fig 3.
User interface of optStoic, requires the molecule information (MetaNetX/KEGG ID or SMILES string) of the starting and target molecules and outputs the optimal overall conversion stoichiometry, i.e., used in novoStoic to find synthesis pathways.
Fig 4.
User interface of novoStoic, requires overall balanced stoichiometry of conversion from a source to a target molecule, and input parameters such as maximum number of steps for pathway search, number of pathways to search, and the number of top known enzyme candidates ranked by EnzRank for novel steps identified in the pathways.
Fig 5.
User interface of dGPredictor, requires the KEGG reaction string as input.
An InChI string is required if a molecule is not present in the database. The user can modify settings such as pH and ionic strength to get at different physiological conditions.
Fig 6.
User interface for EnzRank, requires the SMILES string of the substrate and the enzyme sequence as input to estimate the probability score for the enzyme-substrate compatibility.
Fig 7.
Pathway identified using novoStoic with overall stoichiometry 1, here is the pathway with steps 1-4 showing the known pathway from the literature.
Whereas the pathway with steps 5-3-4 shows a novel pathway that bypasses steps 1 and 2 in a single step. Here, step 1 is a decarboxylation reaction, step 2 is tyramine: oxygen oxidoreductase, step 3 is aldehyde dehydrogenase, step 4 is a hydroxylase reaction, and novel step 5 comes from a reaction MNXR121797 (exact chemical transformation highlighted in blue), which is a hydroxylase enzyme.
Fig 8.
Pathway identified using novoStoic with overall stoichiometry 2, here is the pathway with steps 1-3 showing the shorter novel pathway.
Here, step 1 is a known decarboxylation reaction, step 2 is derived from the reaction rule R2, and step 3 is derived from reaction rule R3.
Table 1.
Top 3 genes from different organisms selected by EnzRank for novel reaction step R3 with reaction rule derived from aminodeoxychorismate synthase enzyme.
Table 2.
Comparison of existing biosynthesis planning tools.