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M200786200v1
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Papers In Press, published online ahead of print March 21, 2002
J. Biol. Chem, 10.1074/jbc.M200786200
Submitted on January 24, 2002
Revised on March 21, 2002
Accepted on March 21, 2002

Isolation and identification of the major heparan sulfate proteoglycans in the developing bovine rib growth plate

Prasanthi Govindraj, Leigh A. West, Thomas J. Koob, Peter Neame, Kurt Doege, and John Hassell

The Center for Research in Skeletal Development and Pediatric Orthopaedics, Shriners Hospitals for Children- Tampa, Tampa, FL 33612

Corresponding Author: jhassell{at}shctampa.usf.edu

Heparan sulfate proteoglycans are thought to mediate the action of growth factors. The heparan sulfate containing proteoglycans in extracts of the bovine fetal rib growth plate were detected using the monoclonal antibody 3G10, which recognizes a neoepitope generated by heparitinase digestion (David, G., et. al, 1992, J. Cell Bio. 119, 961-975). The heparan sulfate proteoglycans that react with this antibody were identified using antisera to known proteoglycans, were purified using CsCl density gradient centrifugation, molecular sieve and ion exchange chromatography and then characterized. The major heparan sulfate proteoglycans in the growth plate had core proteins 200 kD and larger and were identified as perlecan and aggrecan. These two heparan sulfate proteoglycans could be effectively separated from each other by CsCl density gradient centrifugation alone. Perlecan contained 25% heparan sulfate and 75% chondroitin sulfate. The heparan sulfate chains on growth plate perlecan were considerably smaller than the chondroitin sulfate chains and the heparan sulfate disaccharide content was different than that found for heparan sulfate from either kidney, tumor tissue or growth plate aggrecan. Aggrecan contained only 0.1% heparan sulfate which was localized to the CS-1 domain of aggrecan. These results indicate that perlecan and aggrecan would be the principal candidate proteoglycans involved in the action of heparan sulfate binding proteins in the developing growth plate.


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