A flow microcalorimetric method for enzyme activity measurements: application to dihydrofolate reductase.
A flow microcalorimetric method was developed for the analysis of enzymatic activities in crude tissue homogenates. It can be applied whenever a heat exchange is involved in an enzymatic reaction. The consequent sensitivity obviously depends on the enthalpy variation observed. Dihydrofolate reductase was chosen as an example; this enzyme is the molecular target of methotrexate, a widely used anticancer agent. This calorimetric method, whose sensitivity limit is 1.48 X 10(-4) units of dihydrofolate reductase per milliliter of reactant medium, allows enzyme activity measurements in tissues with low dihydrofolate reductase levels. A few examples of measurements in animal tissues are given. These measurements are of some interest; indeed, increased activity and increased levels of this enzyme are two of the mechanisms which may explain resistance to methotrexate.
Sica
L
L
Laboratoire de Physique Pharmaceutique, Marseille, France.
Gilli
R
R
Briand
C
C
Sari
J C
JC
D000818:Animals; D002151:Calorimetry / Q000379:methods; D002417:Cattle; D004798:Enzymes / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008168:Lung / Q000201:enzymology; D051381:Rats; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism*
D004798:Enzymes; D013762:Tetrahydrofolate Dehydrogenase