One-pot enzymatic reaction sequence for the syntheses of d-glyceraldehyde 3-phosphate and l-glycerol 3-phosphate

• One-pot enzymatic reaction sequence was designed for d-GAP and sn-G3P syntheses.
• Reaction sequance overcomes thermodynamic barrier yielding 100% conversion.
• Different reactor types were evaluated and batch-wise operation is most convinenet.
• Separation of d-GAP and sn-G3P has been achieved by polyethylenimine-cellulose TLC.

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.

Figure optionsDownload as PowerPoint slide