STRUCTURAL DOMAINS IN CHONDROITIN SULFATE IDENTIFIED BY ANTI-CHONDROITIN SULFATE MONOCLONAL-ANTIBODIES - IMMUNOSEQUENCING OF CHONDROITIN SULFATES

Monoclonal antibodies have been developed that recognize epitopes in n ative chondroitin sulfate chains. One of these antibodies, CS-56, repo rtedly recognizes chondroitin 4- and 6-sulfates. However, this antibod y, and four other anti-chondroitin sulfate antibodies, 4C3, 4D3, 6C3 a nd 7D4, do not recognize epitopes in chondroitin sulfate chains from S warm rat chondrosarcoma proteoglycan, an indication that native chondr oitin sulfate epitopes are more structurally complex than the standard 0-, 4-, and 6-sulfated disaccharide repeats that constitute the backb one of chondroitin sulfate chains. A series of limited chondroitinase digestions was performed on the large aggregating proteoglycan monomer extracted from embryonic chick chondrocyte cultures to identify the d igestion parameters required to release the different native chondroit in sulfate epitopes. Some epitopes were more accessible to enzymatic d igestion than other epitopes. The approximate location of epitopes was determined by measuring the size of undigested oligosaccharides retai ned on the core protein following a limited digestion, and correlating this with the level of immunoreactivity for the different antibodies. These analyses identified the locations of three different antigenic domains. Domain 1 resides at the linkage region and contains epitopes for two of the five antibodies, and a portion of the epitopes for a th ird antibody. Domain 2 lies in the interior of the chain and contains epitopes for three of the five antibodies. Domain 3 resides at the non -reducing terminus and does not contain epitopes for any of the anti-c hondroitin sulfate antibodies used in this study. These results indica te that specific native chondroitin sulfate epitopes are non-randomly distributed within the linear framework of chondroitin sulfate chains.