Covalent crosslinking analysis of angiotensin receptors on differentiated NG108-15 cells.
Neuroblastoma x glioma hybrid cells (NG108-15) were used as a model system to characterize neuronal-glial type angiotensin (ANG) receptors by covalent crosslinking analysis. After differentiation with 1.5% DMSO and 0.5% fetal bovine serum for four to five days, saturation analysis revealed a single high affinity site with a Kd = 1.35 +/- 0.42 nM and a Bmax = 468 +/- 106 fmol/mg protein. Using the homobifunctional crosslinking reagent bis(sulfosuccinimidyl) suberate (BS3), a site with an estimated Mr of 78 kDa was specifically labeled with 125I-ANG II as determined by SDS-polyacrylamide gel electrophoresis. Both ANG II and ANG III (10(-6) M) inhibited specific labeling. The Ki for ANG III binding was similar by both pharmacologic (Ki = 3.33 +/- 0.98 nM) and gel densitometric (Ki = 2.65 +/- 0.32 nM) analyses. We conclude that the 78 kDa protein represents a high affinity ANG binding site with similar affinities for both ANG II and ANG III.
Carrithers
M D
MD
Neuroscience Program, University of Illinois, Urbana 61801.
Koide
K A
KA
Raman
V K
VK
Masuda
S
S
Weyhenmeyer
J A
JA
D001665:Binding Sites; D002454:Cell Differentiation; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D005910:Glioma / Q000378:metabolism*; D009447:Neuroblastoma / Q000378:metabolism*; D011945:Receptors, Angiotensin / Q000378:metabolism*; D013388:Succinimides / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000378:metabolism
D003432:Cross-Linking Reagents; D011945:Receptors, Angiotensin; D013388:Succinimides; C035760:bis(sulfosuccinimidyl)suberate