Interactions of neurotrophin-3 (NT-3), brain-derived neurotrophic factor (BDNF), and the NT-3.BDNF heterodimer with the extracellular domains of the TrkB and TrkC receptors.
Interactions of three neurotrophin dimers, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and a NT-3.BDNF heterodimer with extracellular, soluble TrkB and TrkC receptors were studied using native gels, light scattering, and sedimentation equilibrium. These three neurotrophins showed binding of two TrkB receptors per neurotrophin dimer, with a tendency to dissociate into one TrkB per dimer for NT-3 and the heterodimer, as determined by native gels, light scattering, and sedimentation equilibrium. For TrkC, native gels suggested binding of NT-3, heterodimer, and BDNF but not of nerve growth factor. Sedimentation equilibrium revealed that all three neurotrophin molecules bind to TrkC at two receptors per dimer but that BDNF binds much more weakly and that the heterodimer has an intermediate binding strength. Light scattering/size exclusion chromatography showed complexes with two TrkC receptors per NT-3 dimer and one TrkC per heterodimer but did not detect binding of BDNF to TrkC. This latter result is not inconsistent with the sedimentation data, because the weak binding of BDNF to TrkC may be easily dissociated by nonspecific interactions of BDNF with the size exclusion column. The relative binding constants for these neurotrophins and the soluble receptor extracellular domains, as determined by sedimentation equilibrium, are correlated with their biological activity. However, the magnitude of these binding constants is insufficient by approximately 3 orders of magnitude to promote receptor dimerization at physiologically active concentrations.
Philo
J
J
Amgen Inc., Thousand Oaks, California 91320-1789.
Talvenheimo
J
J
Wen
J
J
Rosenfeld
R
R
Welcher
A
A
Arakawa
T
T
D000818:Animals; D019208:Brain-Derived Neurotrophic Factor; D016466:CHO Cells; D002850:Chromatography, Gel; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D008024:Ligands; D046911:Macromolecular Substances; D008970:Molecular Weight; D009414:Nerve Growth Factors / Q000302:isolation & purification / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000302:isolation & purification / Q000378:metabolism*; D020933:Neurotrophin 3; D055503:Protein Multimerization; D020794:Receptor Protein-Tyrosine Kinases / Q000302:isolation & purification / Q000378:metabolism*; D020801:Receptor, Ciliary Neurotrophic Factor; D020812:Receptor, trkC; D017475:Receptors, Nerve Growth Factor / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012542:Scattering, Radiation; D014162:Transfection
D019208:Brain-Derived Neurotrophic Factor; D008024:Ligands; D046911:Macromolecular Substances; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D020933:Neurotrophin 3; D020801:Receptor, Ciliary Neurotrophic Factor; D017475:Receptors, Nerve Growth Factor; D011994:Recombinant Proteins; D020794:Receptor Protein-Tyrosine Kinases; D020812:Receptor, trkC