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Activating Silent Glycolysis Bypasses in Escherichia coli

BioDesign Research 2022年第1期2022卷 295-311页

作者： Camillo Iacometti Katharina Marx Maria Hönick Viktoria Biletskaia Helena Schulz-Mirbach Beau Dronsella Ari Satanowski Valérie A.Delmas Anne Berger Ivan Dubois madeleine bouzon Volker Döring Elad Noor Arren Bar-Even Steffen N.Lindner Max Planck Institute of Molecular Plant Physiology Am Mühlenberg 114476 Potsdam-GolmGermany Génomique Métabolique GenoscopeInstitut François JacobCEACNRSUniv EvryUniversitéParis-Saclay91057 Evry-CourcouronneFrance Institute of Molecular Systems Biology ETH ZürichOtto-Stern-Weg 38093 ZürichSwitzerland Department of Plant and Environmental Sciences Weizmann Institute of ScienceRehovotIsrael Department of Biochemistry CharitéUniversitätsmedizinVirchowweg 610117 BerlinGermany

All living organisms share similar reactions within their central metabolism to provide precursors for all essential building blocksand reducing power. To identify whether alternative metabolic routes of glycolysis can operate in E. coli, we complementarilyemployed in silico design, rational engineering, and adaptive laboratory evolution. First, we used a genome-scale model andidentified two potential pathways within the metabolic network of this organism replacing canonical Embden-Meyerhof-Parnas(EMP) glycolysis to convert phosphosugars into organic acids. One of these glycolytic routes proceeds via methylglyoxal andthe other via serine biosynthesis and degradation. Then, we implemented both pathways in E. coli strains harboring defectiveEMP glycolysis. Surprisingly, the pathway via methylglyoxal seemed to immediately operate in a triosephosphate isomerasedeletion strain cultivated on glycerol. By contrast, in a phosphoglycerate kinase deletion strain, the overexpression ofmethylglyoxal synthase was necessary to restore growth of the strain. Furthermore, we engineered the “serine shunt” whichconverts 3-phosphoglycerate via serine biosynthesis and degradation to pyruvate, bypassing an enolase deletion. Finally, toexplore which of these alternatives would emerge by natural selection, we performed an adaptive laboratory evolution studyusing an enolase deletion strain. Our experiments suggest that the evolved mutants use the serine shunt. Our study reveals theflexible repurposing of metabolic pathways to create new metabolite links and rewire central metabolism.

关键词： convert rational sugar

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