Faculty Collaboration Database

Site-directed removal of N-glycosylation sites in the bovine cation-dependent mannose 6-phosphate receptor: effects on ligand binding, intracellular targetting and association with binding immunoglobulin protein. Biochem J 1993 Nov 01;295 ( Pt 3)(Pt 3):841-8

Date

11/01/1993

Pubmed ID

8240300

Pubmed Central ID

PMC1134638

DOI

10.1042/bj2950841

Scopus ID

2-s2.0-0027503915 (requires institutional sign-in at Scopus site)   27 Citations

Abstract

The bovine cation-dependent mannose 6-phosphate receptor (CD-MPR) contains five potential N-linked glycosylation sites, four of which are utilized. To evaluate the function of these oligosaccharides, site-directed mutagenesis was used to generate glycosylation-deficient CD-MPR mutants lacking various potential glycosylation sites. The mutants were constructed in both a full-length and a soluble truncated (STOP155 construct) form of the receptor and their properties were examined in transfected COS-1 cells. The results showed that the presence of a single oligosaccharide chain, particularly at position 87, on the CD-MPR significantly enhanced its mannose 6-phosphate (Man-6-P)-binding ability when compared with non-glycosylated receptors. In addition, the presence of a single oligosaccharide chain at position 87, and to a lesser degree at position 31 or 81, promoted the secretion of the STOP155 CD-MPR. Pulse-labelling of transfected COS-1 cells followed by immunoprecipitation with binding immunoglobulin protein (BiP)-specific and CD-MPR-specific antibodies indicated that BiP associated with the non-glycosylated forms of the receptor but not with the wild-type CD-MPR. Furthermore, the association of the various glycosylation-deficient forms of the CD-MPR with BiP correlated inversely with their ability to bind Man-6-P. From these results we conclude that N-glycosylation of the bovine CD-MPR facilities the folding of the nascent polypeptide chain into a conformation that is conductive for intracellular transport and ligand binding.

Author List

Zhang Y, Dahms NM

Author

Nancy M. Dahms PhD Professor in the Biochemistry department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Base Sequence
Binding Sites
Carrier Proteins
Cattle
Cell Line
Glycosylation
Heat-Shock Proteins
Kidney
Mannosephosphates
Molecular Chaperones
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Conformation
Receptor, IGF Type 2
Transfection