99 research outputs found
From the Particular to the General: Three Federal Rules and the Jurisprudence of the Rehnquist and Roberts Courts
New Approaches to Enforcement and Compliance with Labour Regulatory Standards: The Case of Ontario, Canada
The Analysis of Coordinated Effects in EU Merger Control: Where Do We Stand after Sony/BMG and Impala?
Beilstein-Institut Assay of Enzymes with Insoluble or Unknown Substrates: The Membrane-Bound Quinone
The conventional assay method for the majority of enzymes envisages a reaction between substrates in aqueous solution. A measurable concentration of product is accumulated over time. This paradigm has served well for the characterization of many enzymes. Variations of the method, often using chromogenic or fluorogenic substrates, have been developed and are widely used for purposes such as clinical diagnosis and screening. There are some metabolically important enzymes for which the only published assay methods use artificial substrates. Some of these are oxidoreductases that use artificial mediators, and are listed in the EC list under EC 1.x.99. For computational reconstruction of the metabolism of a cell, however, it is necessary to use kinetic data from assays that reflect the physiological function in the cell, and the physiological substrates. For some oxidoreductases it is known, or considered likely that the acceptors are water-insoluble membrane-bound quinones such as ubiquinone or menaquinone, which present particular problems for measurement of kinetic parameters. Succinate dehydrogenase/fumarate reductase is considered as an example. The oxidoreductases from membranes must be rendered soluble by detergents, which alter their kinetic behaviour. Uncertainty about the way of measuring activity of such enzymes has led to confusion in textbooks and metabolic maps, such as the persistent myth that free FAD is the acceptor for succinate dehydrogenase and related enzymes. New strategies are discussed to measure electrontransfer flux, under conditions that reflect the physiological activity o
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