Characterization of a heparan sulfate proteoglycan that copurifies with the neural cell adhesion molecule.
We have demonstrated previously that the neural cell adhesion molecule (NCAM) interacts with a neuronal heparan sulfate proteoglycan. The binding of this proteoglycan(s) by NCAM appears to be required for NCAM-mediated cell adhesion, although the mechanism is unclear. In the present study we show that a heparan sulfate proteoglycan copurifies with NCAM, and provide an initial biochemical characterization of the proteoglycan. The copurification of a heparan sulfate proteoglycan with NCAM was demonstrated following immunopurification of NCAM from a detergent extract of cell membranes derived from Na2(35)SO4-labeled neural retinal cells. A large-molecular-weight, 35SO4-labeled molecule copurified with NCAM isolated from these neural cell cultures, and was resistant to chondroitinase ABC treatment, but degraded completely by nitrous acid treatment. These results indicate that the molecule is a heparan sulfate proteoglycan. Although this proteoglycan copurifies with NCAM, it is not detected when the neuron-glia cell adhesion molecule (NgCAM) is immunopurified using the 8D9 monoclonal antibody. The heparan sulfate proteoglycan may also be a membrane-associated proteoglycan since it interacts with phenyl-Sepharose. Molecular weight characterization of the proteoglycan by gel filtration chromatography indicates a molecular weight of 400-520 kDa. The heparan sulfate glycosaminoglycan chains were shown to have an average molecular weight of approximately 40 kDa, and the polypeptide backbone was estimated to be 120 kDa by polyacrylamide gel electrophoresis. These data therefore demonstrate that a neuronal heparan sulfate proteoglycan copurifies with NCAM.
Cole
G J
GJ
Department of Anatomy and Cell Biology, Medical University of South Carolina, Charleston 29425.
Burg
M
M
D000818:Animals; D000954:Antigens, Surface / Q000302:isolation & purification* / Q000378:metabolism; D001923:Brain Chemistry; D015815:Cell Adhesion Molecules; D002642:Chick Embryo; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis* / Q000378:metabolism; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005898:Glass; D006025:Glycosaminoglycans / Q000032:analysis*; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate / Q000032:analysis* / Q000378:metabolism; D008970:Molecular Weight; D011485:Protein Binding; D011509:Proteoglycans / Q000032:analysis*; D012160:Retina / Q000032:analysis; D014758:Viral Core Proteins / Q000032:analysis
D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D011508:Chondroitin Sulfate Proteoglycans; D006025:Glycosaminoglycans; D019812:Heparan Sulfate Proteoglycans; D006497:Heparan Sulfate; D011509:Proteoglycans; D014758:Viral Core Proteins