One-pot enzymatic reaction sequence for the syntheses of D-glyceraldehyde 3-phosphate and L-glycerol 3-phosphate

Getachew Shibabaw Molla, Roland Wohlgemuth, Andreas Liese

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A one-pot enzymatic reaction sequence for the synthesis of optically pure D-glyceraldehyde 3-phosphate (D-GAP) and L-glycerol 3-phosphate (sn-G3P) was designed using fructose-bisphosphate aldolase from rabbit muscle (RAMA), sn-glycerol 3-phosphate dehydrogenase (sn-G3PDH) and formate dehydrogenase from Candida boidinii (FDH). The reaction sequence significantly improves the aldol cleavage of D-fructose 1,6-bisphosphate (D-F16BP) catalyzed by RAMA and yields 100% conversion of D-F16BP by overcoming thermodynamic limitation. The degradation kinetics of D-GAP under reaction conditions was investigated and a reaction kinetics model defining the entire cascade was developed. Validation of the model shows 98.5% correlation between experimental data and numerically simulated data matrices. The evaluation of different types of reactor was performed by combining the reaction kinetics model, mass balances and kinetics of the non-enzymatic degradation of D-GAP. Batch-wise operation in a stirred tank reactor (STR) is the most convenient procedure for the one-pot enzymatic syntheses of D-GAP and sn-G3P. The separation of the two products D-GAP and sn-G3P has been achieved using polyethylenimine (PEI)-cellulose TLC.
Original languageEnglish
JournalJournal of Molecular Catalysis B: Enzymatic
Pages (from-to)77-82
Publication statusPublished - 2016
Externally publishedYes


  • Phosphorylated metabolite
  • One-pot reaction sequence
  • Reaction kinetics
  • Reactor simulation
  • Downstream processing

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