Bioluminescent assay of bacterial intracellular AMP, ADP, and ATP with the use of a coimmobilized three-enzyme reagent (adenylate kinase, pyruvate kinase, and firefly luciferase).
A three-enzyme coimmobilized system (firefly luciferase, pyruvate kinase, and adenylate kinase) was constructed for the bioluminescent assay of ATP, ADP, and AMP in bacterial cell extracts. Data for the reproducibility and sensitivity of the proposed method are presented. Detection limits were 1.5 pmol of ADP and 15 pmol of AMP in the sample. With this system, changes in adenine nucleotide concentrations in bacterial cells were measured during the actions exerted by external chemical and physical sources, such as additives to nutrient media and low-power He-Ne laser irradiation.
Chemical Enzymology Department, Chemistry College, Lomonosov Moscow State University, Russia.
Brovko LYu
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Romanova
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Ugarova
D000244:Adenosine Diphosphate / Q000032:analysis*; D000249:Adenosine Monophosphate / Q000032:analysis*; D000255:Adenosine Triphosphate / Q000032:analysis*; D000263:Adenylate Kinase; D000818:Animals; D001419:Bacteria / Q000737:chemistry* / Q000378:metabolism; D001517:Coleoptera; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects; D007834:Lasers; D008156:Luciferases; D008163:Luminescent Measurements; D010101:Oxygen Consumption / Q000528:radiation effects; D011770:Pyruvate Kinase; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D004800:Enzymes, Immobilized; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D008156:Luciferases; D011770:Pyruvate Kinase; D000263:Adenylate Kinase