Characterization of polyphosphate kinases for the synthesis of GSH with ATP regeneration from AMP

Polyphosphate kinase (PPK) is important for industrial processes involving ATP regeneration. While a variety of methods have been reported for regenerating ATP from ADP, few have explored enzyme catalyzed ATP regeneration from cheaper and stable AMP. In this work, PPKs from different sources were expressed and their catalytic activity were tested at different reaction temperatures, reaction pH and with different polyphosphate (polyPn) types. The ATP regeneration system for glutathione (GSH) synthesis was established using a single PPK capable of phosphorylating AMP to synthesize ATP from AMP and short chain polyPn. GSH yield was obtained using adenosine mono-, di- and triphosphates, which confirmed the flexibility of our constructed ATP regeneration system coupled with GSH synthesis via bifunctional GSH synthase. Finally, optimization of the GSH synthesis yielded conversion value above 80 %. Overall, these results illustrate that PPK is suitable for a broader range of substrates than previously expected, and has great untapped potential for applications involving ATP regeneration.

Automated summary

The study demonstrates that PPK plays a crucial role in ATP regeneration in industrial processes. By exploring different sources of PPK, an ATP regeneration system for glutathione synthesis was established, with optimization resulting in a yield above 80%. These findings highlight the versatility of PPK for a wider range of substrates than previously anticipated, showcasing its untapped potential for applications involving ATP regeneration.