Isolation and characterization of insulin receptors from rat kidney glomeruli and tubules.
In order to directly compare the structural characteristics of renal glomerular and tubular insulin receptors, the purified isolated nephron subunits were extracted with 1% Triton X-102, fractionated by DEAE-Sephacel ion exchange column chromatography and the fractions containing insulin binding proteins were identified by the precipitation of 125I-insulin-protein complexes with polyethylene glycol (PEG). The fractions containing insulin binding proteins were pooled, incubated with 125I-insulin and covalently cross-linked with disuccinimidyl suberate, followed by chromatography of the cross-linked samples on Sepharose CL-6B. From both glomeruli and tubules, three 125I-insulin-binding complexes with molecular weights of 560 KDa, 220 KDa and 95 KDa were found. SDS-PAGE of these complexes from glomeruli and tubules under both reducing and nonreducing conditions gave similar patterns of 125I-insulin-crosslinked components, with the exception of the polypeptide pattern from the 560 KDa peak fraction which was markedly different between glomeruli and tubules with the former giving major labeled components at 170 and 68 KDa while the latter showed labeled components of 125 KDa and greater than 250 KDa. Glomerular and tubular insulin receptors, therefore, display similar subunit composition under reducing conditions, but differ in the non-reduced state, suggesting that these complexes may differ in the extent and/or nature of disulfide bonding.
Im
J H
JH
Department of Pharmacology, University of Alabama, Birmingham 35294.
Pillion
D J
DJ
Meezan
E
E
D000818:Animals; D001345:Autoradiography; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007678:Kidney Glomerulus / Q000032:analysis* / Q000648:ultrastructure; D007684:Kidney Tubules / Q000032:analysis* / Q000648:ultrastructure; D008970:Molecular Weight; D051381:Rats; D011972:Receptor, Insulin / Q000032:analysis / Q000302:isolation & purification*
D011972:Receptor, Insulin