Characterization of an epithelial similar to 460-kDa protein that facilitates endocytosis of intrinsic factor-vitamin B-12 and binds receptor-associated protein

被引:114
作者
Birn, H
Verroust, PJ
Nexo, E
Hager, H
Jacobsen, C
Christensen, EI
Moestrup, SK
机构
[1] AARHUS UNIV,DEPT BIOCHEM MED,DK-8000 AARHUS C,DENMARK
[2] AARHUS UNIV,DEPT CELL BIOL,DK-8000 AARHUS C,DENMARK
[3] AARHUS KOMMUNE HOSP,DEPT CLIN BIOCHEM,DK-8000 AARHUS C,DENMARK
[4] HOP TENON,INSERM,U64,F-75020 PARIS,FRANCE
关键词
D O I
10.1074/jbc.272.42.26497
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
By using receptor-associated protein (RAP) as an affinity target, an intrinsic factor-vitamin B-12 (IF-B-12)-binding renal epithelial protein of similar to 460 kDa was copurified together with the transcobalamin-B-12-binding 600-kDa receptor, megalin, IF-B-12 affinity chromatography of renal cortex membrane from rabbit and man yielded the same similar to 460-kDa protein. Binding studies including surface plasmon resonance analyses of the protein demonstrated a calcium-dependent and high affinity binding of IF-B-12 to a site distinct from the RAP binding site. The high affinity binding of IF-B-12 was dependent on complex formation with vitamin B-12. Light and electron microscope autoradiography of rat renal cortex cryosections incubated directly with IF-Co-57-B-12 and rat proximal tubules microinjected in vivo with the radioligand demonstrated binding of the ligand to endocytic invaginations of proximal tubule membranes followed by endocytosis and targeting of vitamin B-12 to lysosomes. Polyclonal antibodies recognizing the similar to 460-kDa receptor inhibited the uptake, Immunohistochemistry of kidney and intestine showed colocalization of the IF-B-12 receptor and megalin in both tissues. In conclusion, we have identified the epithelial IF-B-12-binding receptor as a similar to 460-kDa RAP-binding protein facilitating endocytosis.
引用
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页码:26497 / 26504
页数:8
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