A radioactive binding assay for inhibitors of trkA kinase

Analytical Biochemistry
E KnightC A Dionne

Abstract

The high-affinity receptor for nerve growth factor (NGF), trkA, is a receptor-linked tyrosine kinase. The binding of NGF to trkA, depending on the context of its environment, can cause beneficial or deleterious responses in the target cells. For example, the activation of trkA in sympathetic and sensory neurons causes the subsequent survival and differentiation of these cells. On the other hand, the activation of trkA by NGF in other cells has been implicated in several pathologies including inflammation-induced hyperalgesia and several cancers. A radioactive binding assay to evaluate inhibitors of the kinase domain of trkA has been developed and validated. The assay monitors the specific binding of an inhibitor of trkA kinase activity, the indolocarbazole K-252a, to the trkA receptor. [3H]K-252a binds with high affinity to one site on the cytoplasmic kinase domain of the trkA receptor. Binding is saturable and reversible with a dissociation constant (Kd) of 1.5 nM. The binding assay has been used in competition binding experiments to determine the inhibition constants for other indolocarbazole compounds. The IC50 values for compounds obtained in the binding assay correlate very well with the IC50 values obtained in an enzyme-l...Continue Reading

References

Nov 27, 1991·Biochemical and Biophysical Research Communications·Y HashimotoS Nakanishi
Aug 31, 1990·Biochemical and Biophysical Research Communications·L H ElliottJ S Nixon
Jan 30, 1987·Biochemical and Biophysical Research Communications·H KaseM Kaneko
Dec 1, 1994·The European Journal of Neuroscience·G R LewinL M Mendell
Jun 15, 1995·Biochemical and Biophysical Research Communications·E KnightN Neff
Apr 5, 1996·Analytical Biochemistry·T S AngelesC A Dionne

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Citations

Dec 9, 1998·Proceedings of the National Academy of Sciences of the United States of America·H MorenoR Llinás
Jul 9, 2016·Behavioural Pharmacology·Renata C Dos ReisJuliana G Chichorro

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