Kinetics of a self-amplifying substrate cycle: ADP-ATP cycling assay.
A kinetic study of an ATP-ADP amplification cyclic system involving the enzymes adenylate kinase, pyruvate kinase and L-lactate dehydrogenase has been made. The stoichiometry of the cycle is 2:1, because two molecules of ADP are synthesized from one each of ATP and AMP, and one molecule of ADP is converted back into one of ATP at each turn of the cycle. This results in a continuous exponential increase in the concentrations of ATP and ADP in the reaction medium, according to the equations obtained. This is therefore a substrate cycle that amplifies itself, the cycling rate increasing continuously with time. The background signal of the reagent was reduced by using apyrase to degrade ATP and ADP in the reagent, permitting detection limits as low as 16 pmol of ATP and/or ADP in a continuous spectrophotometric assay.
Departamento de Química-Física, Escuela Politécnica Superior de Albacete, Universidad de Castilla-La Mancha, Campus Universitario, E-02071 Albacete, Spain. evalero@pol-ab.uclm.es
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E
Valero
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R
Varón
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F
García-Carmona
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000263:Adenylate Kinase / Q000378:metabolism; D007700:Kinetics; D011770:Pyruvate Kinase / Q000378:metabolism
D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D011770:Pyruvate Kinase; D000263:Adenylate Kinase