Abstract:
:We identified a 45 kDa Ca(2+)-dependent Lf binding protein on rat hepatocytes. Dithiobis(sulfosuccimidylproprionate) (DTSSP)-crosslinked 125I-Lf to a 45 kDa adduct in a Ca(2+)-dependent manner on intact cells. The 125I-labeled crosslinked complexes were absent when either surface-bound 125I-Lf was stripped prior to crosslinking or an excess of unlabeled Lf was included in the DTSSP reaction. Triton X-100 extracts of hepatocyte membrane ghosts were chromatographed on Lf-agarose, and a 45 kDa polypeptide (p45) was eluted by EGTA. Anti-p45 sera blocked vigorously 125I-Lf endocytosis to intact rat hepatocytes, confirming that p45 functions as the Ca(2+)-dependent Lf receptor on hepatocytes. Two tryptic fragments of p45 showed 100% identity with internal sequences (Leu121-->Lys126 and Phe198-->Lys220) of the major subunit (RHL-1) of the rat asialoglycoprotein receptor. Antisera against p45 and RHL-1 crossreacted equally well with each protein, and asialoorosomucoid blocked the binding of 125I-Lf to hepatocytes. We did not detect the minor subunits (RHL-2/3) of the rat asialoglycoprotein receptor in p45 preparations from Triton X-100-extracts of hepatocytes, and 125I-Lf bound to immobilized RHL-1 but not to RHL-2/3. Exoglycosidases were used to remove terminally-exposed NeuNAc and alpha- and beta-Gal from bovine Lf glycans, and lectin blotting confirmed that glycosidase-treated Lfs lacked detectable terminal Gal. Unexpectedly, deglycosylated Lf exhibited no loss in its ability to compete with unmodified Lf for binding to isolated hepatocytes. Moreover, beta-lactose but not sucrose competed vigorously for 125I-Lf endocytosis by hepatocytes, indicating that Lf binds at or near the carbohydrate-recognition domain of RHL-1. We conclude that RHL-1 is the Ca(2+)-dependent Lf receptor on hepatocytes and that it binds Lf in a Gal-independent manner.
journal_name
Adv Exp Med Bioljournal_title
Advances in experimental medicine and biologyauthors
McAbee DD,Bennatt DJ,Ling YYsubject
Has Abstractpub_date
1998-01-01 00:00:00pages
113-21eissn
0065-2598issn
2214-8019journal_volume
443pub_type
杂志文章abstract::Human genetic factors play an important role in determining the outcome of infections caused by intracellular pathogens, including mycobacteria and salmonellae (reviewed in 1). The genetic elements involved and the mechanisms by which these control disease-susceptibility versus resistance, however, remain incompletely...
journal_title:Advances in experimental medicine and biology
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doi:10.1007/978-1-4615-0059-9_24
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journal_title:Advances in experimental medicine and biology
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journal_title:Advances in experimental medicine and biology
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journal_title:Advances in experimental medicine and biology
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更新日期:2016-01-01 00:00:00
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journal_title:Advances in experimental medicine and biology
pub_type: 杂志文章,评审
doi:10.1007/978-981-10-5562-1_6
更新日期:2017-01-01 00:00:00
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