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|Background:||The Trk (tropomyosin receptor kinase) receptors belong to the family of receptor tyrosine kinases, and three trk genes have been identified in mammals. The TrkA protooncogene was first identified as a nerve growth factor receptor (NGFR) (1, 2), followed by the identification of TrkB and TrkC (3). These Trk receptors are transmembrane glycoproteins of ~140 kDa. They are tyrosine kinases with an extracellular ligand-binding domain containing multiple repeats of leucine-rich motifs, two cysteine clusters (C1, C2), two immunoglobulin-like domains (Ig1, Ig2), and a single transmembrane domain (4). The tyrosine kinase domains are highly related (~80% amino acid identity), however the extracellular domains are more divergent (~30% identity). Nerve growth factor (NGF) is the preferred ligand for TrkA, brain-derived neurotrophic factor (BDNF) and Neurotrophin-4/5 (NT-4/5) are preferred for TrkB, and Neurotrophin-3 (NT-3) for TrkC (3). These specificities are not absolute, and NT-3 is also a ligand for TrkA and TrkB. Nerve growth factor (NGF) is the preferred ligand for TrkA, brain-derived neurotrophic factor (BDNF) and Neurotrophin-4/5 (NT-4/5) are preferred for TrkB, and Neurotrophin-3 (NT-3) for TrkC (3). These specificities are not absolute, and NT-3 is also a ligand for TrkA and TrkB. Neurotrophin binding to Trk receptors triggers dimerization leading to the activation of different signaling pathways through recruitment of various adapter molecules. The main binding sites for Trk substrates are two tyrosine residues that, on activation of the Trk receptors, become phosphorylated. Two complexes of adapter molecules bind to the tyrosine residue located in the juxtamembrane region of the Trk receptor, the Shc/Grb2/SOS and the FRS2/SHP-2/Grb2/SOS complex. Ras/Raf/MEK/MAPK induces the differentiation of neurons and neurite growth (5). Measurement of TrkA Kinase activity The protocol generally regarded as most sensitive for the quantitative measurement of TrkA kinase activity involves incubation of the TrkA kinase sample with substrate (either a natural or synthetic polypeptide such as poly[Glu, Tyr]4:1), in the presence of Mg2+, Mn2+ and 32P-labeled ATP. The reaction is terminated by "spotting" a sample onto a filter paper disc, followed by immersion in acid to precipitate the radiolabeled product. The filter papers are then washed extensively to remove unincorporated radiolabel and the radioactivity is counted. While sensitive, this method is labor-intensive, generates hazardous radioactive waste and depends on a radioisotope with a short half-life. It is particularly unsuitable when kinase assays are only performed on an infrequent basis. The CycLexÂ® TrkA Kinase Assay/Inhibitor Screening Kit uses a horseradish peroxidase coupled anti-phosphotyrosine monoclonal antibody as a reporter molecule in a 96-well ELISA format. This assay provides a non-isotopic, sensitive and specific method to detect kinase activity of the TrkA catalytic domain.|
|Gene ID Human:||4914|
|Gene ID Mouse:||18211|
|Regulatory Statement:||For Research Use Only. Not for use in diagnostic procedures.|
There are no references for CycLex TrkA Kinase Assay/Inhibitor Screening Kit at this time.