Role of the oligosaccharide inner core in the inhibition of human leukocyte elastase by chondroitin sulfates.

A chondroitin 4-sulfate and 6-sulfate mixture (CS) extracted from selachian cartilage and a purified preparation of the above glycosaminoglycans from bovine trachea, have been comparatively assayed with respect to the kinetics of human elastase activity. The selachian extract CS, but not the bovine one, inhibited the hydrolysis of N-tert-butyloxycarbonyl p-nitrophenyl ester in vitro (ki = 4.0 micrograms/ml). By fractioning both extracts on Sephadex, the inhibition was found to be related to the presence of a 40-50 KD peak (ki = 2.3 micrograms/ml). This particular fraction was present in the selachian extract CS from matrix, being absent from the bovine trachea extract CS (70% CSC) from Sigma. It showed a single Coomassie- and PAS-stained band approximately 47 kDa in m.w. on sodium dodecylsulfate polyacrylamide gel electrophoresis. Cleavage of the O-glycoside link by alkali hydrolysis of the eluted band, did not influence the enzyme inhibition (ki = 2.1 micrograms/ml). At variance, extensive hydrolysis of ester linkages by fluoride ion resulted in the lowering of inhibition (ki = 5.8 micrograms/ml). Apparently, the preservation of the trisaccharide region in the carbohydrate residues of proteoglycan is mandatory for the inhibition of elastase by low-sulfated chondroitinsulfates.