Detection and characterization of heparin-binding proteins with a gel overlay procedure. |
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Authors: | T Mehlman W H Burgess |
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Institution: | Jerome H. Holland Laboratory for Biomedical Research, Laboratory of Molecular Biology, American Red Cross, Rockville, Maryland 20850. |
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Abstract: | The binding of 125I-labeled derivatives of heparin has been used by several investigators to identify heparin-binding fragments of different heparin-binding proteins. In this report we utilize the procedure described by J.W. Smith and D.J. Knauer (1987, Anal. Biochem. 160, 105-114) to produce 125I-fluorescein-heparin. Using this derivative, we compare the use of gel overlay procedures with "Western blot" procedures for the detection of heparin-binding proteins following polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. We show that the gel overlay procedure is a relatively simple and sensitive method for visualizing heparin-binding proteins. In addition, we use the procedure to characterize the heparin-binding properties of heparin-binding growth factor 1 (acidic fibroblast growth factor) with synthetic peptide competitors and site-directed mutants of the growth factor. |
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