1. Unbalanced Reactions by Markus Krummenacker Q1 2015

2. FBA needs balanced reactions If an FBA model contains unbalanced reactions, it can cause infinite sources of material Thus, reactions have to balance all the atoms on the left with the atoms on the right Including H+ /charges. Our convention is compounds at pH7.3 Reaction balance checker filters away reactions that are unbalanced or undetermined

3. .log file lists unbalanced reactions Lists unbalanced reactions for the current PGDB (but not for MetaCyc) Several sections of problematic reactions: – In pathways – In signalling-pathways (which contain non-metabolic protein metabolites) – Standalone small-molecule reactions – All other problematic reactions. Usually, macromolecule metabolites. Usually, not involved in core metabolism. – Without a path from the nutrients

4. Common sources of problems (1) SRI has ongoing curation efforts to fix erroneous reactions. By now, most straight forward errors in compound structures or reaction equations have been fixed. Sometimes, in old PGDBs, reactions are wrong, which no longer exist in MetaCyc, and so they can’t be updated automatically. Some reactions describe fairly unknown biochemistry. Lack of knowledge is indicated by the CANNOT-BALANCE? flag (in Reaction Editor).

5. Common sources of problems (2) H+ unbalanced, with metabolite classes. – Protonation by Marvin can not handle structures with R groups. Thus generic reactions have not been automatically proton-balanced. – Have to be manually balanced by using Marvin to edit structure (by educated guess) Class metabolites without structures. – Macromolecule metabolites, like redox proteins – Tricky class metabolites for extremely generic classes. Example: SUGAR-PHOSPHATASE-RXN

6. Remedies Add/Edit metabolite structures with Marvin Edit reaction with Reaction Editor – Balance checker in editor, shows residual Special cases of the balance checker: – Structure with R groups: R, R1, R2, R3, R4, R5 – MOLECULAR-WEIGHT slot set (by Frame Editor) to less than 0.01 (e.g. E-, light) – Frame pairs, where one is a modified form of the other. Structure of modified form has to use unmodified frame as an “atom”. Only 1 such reference supported currently. Example: NAD(P)+ / NAD(P)H

7. Consistency Checker Under Manual Tasks, “Generate reaction balance report” Independent of FBA. But a good idea in any case. Right-click on listed reaction, to show or fix

8. Compartments of Transporters For FBA modelling core metabolism, a few transport reactions are essential: ATP synthase, some ETRs like cytochrome oxidase, NADH:ubiquinone oxidoreductase Pathologic inference for a PGDB may not yet infer the correct compartments. RXN-LOCATIONS slot will be cleared, and so defaults are “in” and “out” Reaction Editor can be used to manually fix compartment assignments