1: Introduction
    To increase the number of value-added chemicals that can be produced by metabolic engineering and synthetic biology, constructing metabolic space with novel reactions is very necessary. However, with a large amount of reactions contained in the metabolic space and complicated metabolism within hosts, it is an arduous task to identify novel-pathways linking two molecules or heterologous pathways when engineering a host to produce target. Hence, we built a user-friendly webserver – novoPathFinder, which has several features. (1): It not only allows to design novel-pathways for production of target from specified precursor, but also could identify heterologous novel-pathways when engineering E. coli or Yeast for production of target without providing precursor.; (2): It supports calculation of overall stoichiometric conversions, theoretical yield and thermodynamic feasibility, the last two could be calculated in real-time under customized physical conditions; (3): It could rank pathways with considering steps length, enzyme promiscuity and SCScore. According to the results, it not only could identify experimental pathways in many cases which current webserver tools couldn't recover, but also could predict more efficient solutions.

2: Three functional modules in novoPathFinder


    (1) No-hosts module: novoPathFinder allows to design novel-pathways for production of target from specified precursor without considering hosts.
    (2) E.coli-based module: This module allows users to design heterologous novel-pathways for production of target molecule when engineering E.coli.
    (3) Yeast-based module: This module allows users to design heterologous novel-pathways for production of target molecule when engineering Yeast.

3:  Query Submitted -- E. coli-based module

    (1) Target Name: Input a target name;
    (2) Step(s): It must be an integer, which means that how many heterologous reactions involved in candidate pathway at most. Of course, the computing time will be prolonged with its increasment,the maximum number of heterologous reactions is limited to 10 for the sake of computation time.
    (3) Iterations: Because the retrosynthesis algorithm utilized in novoPathFinder is iterative, users should enter the iteration-searching times. The retrosynthesis algorithm proceeds until the customized iterations are reached, and each iteration will stop until the predefined steps or the precursor/sink metabolites are reached. For example: 1,000 times or more. Of course, the computation time will be prolonged with its increasment.

4: Pathway enumeration



5: Pathway detail


    (1) Main Carbon Resource/Condition : Main Carbon Resource and Condition allow users to specify organism growth condition(main carbon source/oxygen state).
    (2) Biomass: It represents a percentage that minimum growth rate of mutant type accounts for maximum theoretical growth rate of WT type. Users could enter a decimal between 0 and 1.
    * Theoretical yield and carbon fluxes are calculated by using method from FBA[1].
6: Reaction detail


    (1) Reference Rxn: A known reaction from which the chemical transformation are derived.
    (2) Reference Flag: It is represented by yes or no, in which yes means that the reaction is known, and vice versa.
    (3) Penalty Score of enzyme promiscuity: The penalty score of a known reaction was set to 0, otherwise, it was calculated by using the method proposed by Retropath 2.0 [2].
    (4) Thermodynamic feasibility: Thermodynamic feasibility of each reaction is represented by ΔG'm, of which negative value means that the reaction is thermodynamically favorable, and vice versa. ΔG'm could be calculated in real time by using equilibrator-api [3] under customized ph, ionic strength, and temperature. Moreover, we used the 1 mM concentration for all reactants


7: Reference:
        [1].What is flux balance analysis? Jeffrey D Orth, Ines Thiele & Bernhard Ø Palsson.NATURE BIOTECHNOLOGY.2010
        [2].RetroPath2.0: A retrosynthesis workflow for metabolic engineers.Metabolic Engineering.2018
        [3].Consistent estimation of Gibbs energy using component contributions. Noor E, Haraldsdóttir HS, Milo R, Fleming RMT.PLoS Comput Biol.2013
8: Contact us
        Send your comments or error report to RxnFinder Team.
        Email: 164362170@qq.com