A kinetic method for the detection of inhibitory contaminants in radioactive substrates |
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| Authors: | Keith F. Tipton Timothy J. Mantle Miles D. Houslay |
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| Affiliation: | 1. Department of Biochemistry, Tennis Court Road, Cambridge CB2 1QW, England;2. Department of Biochemistry, University of Manchester Institute of Science and Technology, P.O. Box 88, Manchester M6Q 1QD, England |
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| Abstract: | A simple kinetic test is proposed that will detect the presence of contaminating inhibitors in the radioactive substrate solutions used in the radiochemical assays of enzymes. Double-reciprocal plots are constructed from initial velocity data obtained from assays conducted under conditions of constant specific radioactivity and of constant radioactivity. A significant difference between the Km values determined in this way will show the presence of an inhibitory impurity necessitating the purification of the radioactive substrate. It is possible to determine whether the contaminating inhibitor is competitive, uncompetitive or mixed by this method and to estimate the true Km value in the first two cases. |
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