Hydrolytic approach for production of deoxyribonucleoside- and ribonucleoside-5 -monophosphates and enzymatic synthesis of their polyphosphates.
A new, simple, and ingenious method for enzymatic synthesis of deoxy- and ribonucleoside-5 -triphosphates (dNTP and NTP, respectively) has been developed. The method includes the following stages: hydrolysis of DNA with DNase and immobilized S1-nuclease, phosphorylation of the resulting deoxy- and ribonucleoside-5 -monophosphates (dNMP and NMP, respectively) with nucleotidyl kinase from Escherichia coli, and purification by chromatography of the synthesized dNTP and NTP. dNMP was phosphorylated using an ATP-regenerating system based on acetokinase from E. coli and lithium acetylphosphate.
Bochkov
D V
DV
Novosibirsk Pedagogical University, Novosibirsk, Russia. bochkov@nioch.nsc.ru
Khomov
V V
VV
Tolstikova
T G
TG
D000084:Acetate Kinase / Q000302:isolation & purification / Q000378:metabolism; D000818:Animals; D019149:Bioreactors; D004247:DNA / Q000378:metabolism; D003854:Deoxyribonucleotides / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000302:isolation & purification / Q000378:metabolism; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000201:enzymology; D006868:Hydrolysis; D009713:Nucleotidyltransferases / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D012313:RNA; D012265:Ribonucleotides / Q000378:metabolism*; D012474:Salmon; D017640:Silicates; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism
D003854:Deoxyribonucleotides; D004800:Enzymes, Immobilized; D012265:Ribonucleotides; D017640:Silicates; D012313:RNA; D004247:DNA; D000084:Acetate Kinase; D009713:Nucleotidyltransferases; D004706:Endodeoxyribonucleases; D015719:Single-Strand Specific DNA and RNA Endonucleases