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A conserved phosphatase destroys toxic glycolytic side products in mammals and yeast

Abstract

Metabolic enzymes are very specific. However, most of them show weak side activities toward compounds that are structurally related to their physiological substrates, thereby producing side products that may be toxic. In some cases, 'metabolite repair enzymes' eliminating side products have been identified. We show that mammalian glyceraldehyde 3-phosphate dehydrogenase and pyruvate kinase, two core glycolytic enzymes, produce 4-phosphoerythronate and 2-phospho-L-lactate, respectively. 4-Phosphoerythronate strongly inhibits an enzyme of the pentose phosphate pathway, whereas 2-phospho-L-lactate inhibits the enzyme producing the glycolytic activator fructose 2,6-bisphosphate. We discovered that a single, widely conserved enzyme, known as phosphoglycolate phosphatase (PGP) in mammals, dephosphorylates both 4-phosphoerythronate and 2-phospho-L-lactate, thereby preventing a block in the pentose phosphate pathway and glycolysis. Its yeast ortholog, Pho13, similarly dephosphorylates 4-phosphoerythronate and 2-phosphoglycolate, a side product of pyruvate kinase. Our work illustrates how metabolite repair enzymes can make up for the limited specificity of metabolic enzymes and permit high flux in central metabolic pathways.

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Figure 1: Elimination of the GAPDH side product 4-P-erythronate by PGP.
Figure 2: Accumulation of 4-P-erythronate in PGP KO cells leads to inhibition of the pentose phosphate pathway.
Figure 3: 2-P-lactate accumulating in PGP KO cells can inhibit the production of fructose-2,6-BP.
Figure 4: The metabolite repair system is conserved in S. cerevisiae.
Figure 5: 4-P-erythronate is formed by GAPDH via a 1,4-BP-erythronate intermediate, which is dephosphorylated by acylphosphatase or phosphoglycerate kinase.
Figure 6: Schematic representation of the metabolite repair function of PGP in central carbon metabolism.

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Acknowledgements

Funding was provided from WELBIO (CR-2015A-09 to E.V.S.), the Belgian National Science Fund (T.0120.14 to E.V.S. and J.0044.14 to G.T.B.), the Training Fund for Research in Industry and Agriculture (to F.B. and J.B.), the Belgian Cancer Foundation (2010-155 to G.T.B. and 2014-298 to E.V.S.), the Interuniversity Attraction Pole (IAP-P7.43 to E.V.S.), the European Union Seventh Framework Programme (FP7/2007–2013 grant no. 276814 to C.L.L. and EURO-CDG to E.V.S.) and Horizon 2020 (E-RARE-3: EURO-CDG2 to E.V.S.), the de Duve Institute and the Université Catholique de Louvain. We would like to thank E.R. Fearon (University of Michigan), F. Zhang (Massachusetts Institute of Technology), V. Delcenserie (University of Liège), G. Daube (University of Liège), L. Novellasdemunt and R. Bartrons (University of Barcelona) for reagents, R. Gemayel (Catholic University of Leuven) for help with yeast experiments, C. Jäger and C. Singh (University of Luxembourg) for help with the GC/MS analysis, P. Sonveaux (Université Catholique de Louvain) and P.E. Porporato (Université Catholique de Louvain) for help with the Seahorse XF bioanalyzer (grant F.R.S.-FNRS FRFC 2.5025.12) and A.D. Hanson, L. Hue, J.-F. Collet and A. Peracchi for critical reading of the manuscript.

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The study was mainly designed and written by G.T.B., E.V.S., F.C. and F.B. G.T.B. and E.V.S. supervised the work and are equally contributing corresponding authors. All authors contributed to the interpretation of the results, participated in the writing of the manuscript and approved the final version. F.C. identified and purified PGP and ACYP1. F.C., F.B. and I.G. performed enzymatic analysis. F.B., I.G., G.T.B., J.G. and C.L.L. performed GC/MS analysis. G.T.B., I.G., J.B., M.V. and F.B. generated and analyzed mammalian cell lines. J.B. measured oxygen consumption rate. F.B. generated and analyzed yeast strains. G.N., E.V.S. and I.G. performed PFK-2 analysis. A.H. and M.H.R. cloned and produced PFKFB proteins. D.V. and V.S. performed MS analysis.

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Correspondence to Emile Van Schaftingen or Guido T Bommer.

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Supplementary Results, Supplementary Figures 1–15 and Supplementary Tables 1–3. (PDF 3394 kb)

Supplementary Data Set 1

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Supplementary Data Set 2

Raw GC-MS data (XLSX 157 kb)

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Collard, F., Baldin, F., Gerin, I. et al. A conserved phosphatase destroys toxic glycolytic side products in mammals and yeast. Nat Chem Biol 12, 601–607 (2016). https://doi.org/10.1038/nchembio.2104

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