Self-assembled immobilized polyphosphate kinase for ATP and GTP synthesis
Polyphosphate kinase(PPK)can utilize widely distributed,cost-effective,and stable polyphosphate as phosphate donors for the synthesis of ATP and GTP.The latter provides essential energy for cells and participates in the synthesis of numerous phosphate-containing important compounds.This study aimed to utilize new synthetic biology techniques to efficiently and inexpensively synthesize ATP and GTP through polyphosphate kinase catalysis.Firstly,based on the analysis of the catalytic capabilities of different PPK families towards various substrates,DrPPK2 belonging to the PPK family 2 and originating from Deinococcus radiodurans was selected.Its activity in both synthesizing ATP and GTP was confirmed.Subsequently,optimization of pH,Mg2+in the buffer,and temperature significantly im-proved the efficiency of GTP synthesis,with a maximum conversion rate exceeding 92％.To simplify the catalytic process and reduce pro-duction costs,the self-assembling tag CipA was added to achieve the self-assembly immobilization of DrPPK2.High-purity protein could be obtained through centrifugation,enabling enzyme reuse with a conversion rate greater than 90％,and an effective repetition efficiency ex-ceeding 10 times.This study provides a convenient,efficient,and easily scalable self-assembling immobilization system for ATP and GTP biosynthesis,offering insights into the synthesis of other phosphate-containing compounds.
polyphosphate kinaseATPGTPoptimization of catalytic systemself-assembled immobilized
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维普
