Identification of a 175 kDa protein as the ligand-binding subunit of the rat liver sinusoidal endothelial cell hyaluronan receptor |
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Authors: | Yannariello-Brown Judith; Zhou Bin; Weigel Paul H |
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Institution: | Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center Oklahoma City, OK 73190, USA
1Departments of Ophthalmology & Visual Sciences, and Human Biological Chemistiy & Genetics, University of Texas Medical Branch, Galveston, TX 77555-0787, USA |
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Abstract: | The rat liver sinusoidal endothelial cell (LEC) hyaluronan (HA)receptor was previously identified using a photoaffinity HAderivative (J. BioL Chem., 267, 2045120456, 1992). Twopolypeptides with Mr = 175,000 and 166,000, were consistentlycrosslinked, suggesting that the LEC HA receptor is an oligomer.Whether one or both subunits participate in HA binding, wasnot determined. Here we investigate the HA-subunit interactionsand the potential oligomeric nature of the LEC HA receptor.When Sephacryl-400 gel filtration chromatography was used toenrich the HA receptor, the 175 kDa polypeptide was the majorband seen by SDS-PAGE analysis. Little staining was seen at166 kDa, suggesting that the 175 kDa protein could be separatedfrom the 166 kDa protein and still retain HA-binding activity.A ligand blot assay was used to determine if each individualsubunit could bind HA. LEC proteins were separated by nonreducingSDS-PAGE, and then immobilized onto nitrocellulose. 125I-HAbound to a 175 kDa polypeptide but not to the 166 kDa protein.A high molecular weight band of |
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