Human stem cell factor dimer forms a complex with two molecules of the extracellular domain of its receptor, Kit.
Stem cell factor (SCF) is a cytokine that is active toward hematopoietic progenitor cells and other cell types, including germ cells, melanocytes, and mast cells, which express its receptor, the tyrosine kinase, Kit. SCF exists as noncovalently associated dimer at concentrations where it has been possible to study its quaternary structure; it stimulates dimerization and autophosphorylation of Kit at the cell surface. We have used recombinant versions of human SCF and human Kit extracellular domain (sKit) to study SCF-Kit interactions. By size exclusion chromatography, plus various physical chemical methods including light scattering, sedimentation equilibrium, and titration calorimetry, we demonstrate the formation of complexes containing a dimer of SCF (unglycosylated SCF1-165) plus two molecules of sKit. The concentrations of SCF and sKit in these studies were in the range of 0.35-16.2 microM. The data are analyzed and discussed in the context of several possible models for complex formation. In particular, the sedimentation data are not consistent with a model involving cooperative binding. The Kd estimate for SCF-sKit interaction, obtained by sedimentation equilibrium, is about 17 nm at 25 degrees C. With glycosylated SCF1-165, the Kd is considerably higher.
Philo
J S
JS
Amgen Inc., Thousand Oaks, California 91320, USA.
Wen
J
J
Wypych
J
J
Schwartz
M G
MG
Mendiaz
E A
EA
Langley
K E
KE
D000818:Animals; D016466:CHO Cells; D002151:Calorimetry; D002850:Chromatography, Gel; D006224:Cricetinae; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008027:Light; D019009:Proto-Oncogene Proteins c-kit / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D012542:Scattering, Radiation; D019089:Stem Cell Factor / Q000737:chemistry* / Q000235:genetics
D011994:Recombinant Proteins; D019089:Stem Cell Factor; D019009:Proto-Oncogene Proteins c-kit