Detection of the basic fibroblast growth factor low affinity binding site in cardiac sarcolemmal vesicles.
Bovine cardiac sarcolemmal (SL) vesicles contain basic fibroblast growth factor (bFGF)-binding proteins. Binding to native SL vesicles was specific and saturable with a Kd of 6.9 nM and a Bmax of 15.2 pmoles bFGF/mg vesicle protein. Using radioiodinated bFGF as a probe, autoradiography of SL proteins subjected to SDS-PAGE and electroblotting onto nitrocellulose revealed a set of 3-4 bands, of an apparent molecular weight of 100-150 kDa. bFGF binding to these bands was reduced by pretreatment of SL vesicles with heparinase. Binding was abolished by treatment of blot strips with heparinase or high salt concentrations (greater than 0.6 NaCl) but not endoglycosidase F. bFGF-binding activity remained associated with the membrane fraction following an alkaline wash, which removed peripheral membrane proteins. These data suggest that the cardiac SL contains an integral proteoglycan(s) which may be a low affinity binding/storage site of endogenous bFGF.
Ross
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CR
Dalton Research Center, Department of Veterinary Biomedical Sciences, University of Missouri, Columbia, MO 65211.
Hale
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CC
D000818:Animals; D001345:Autoradiography; D001665:Binding Sites; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism; D019763:Heparin Lyase; D015151:Immunoblotting; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D009206:Myocardium / Q000378:metabolism*; D011133:Polysaccharide-Lyases / Q000378:metabolism; D011509:Proteoglycans / Q000032:analysis* / Q000378:metabolism; D012508:Sarcolemma / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
D011509:Proteoglycans; D012965:Sodium Chloride; D005346:Fibroblast Growth Factors; D006026:Glycoside Hydrolases; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase