DIFFERENTIATION OF CONGLUTINATION ACTIVITY AND SUGAR-BINDING ACTIVITY OF CONGLUTININ AFTER REMOVAL OF NH2-TERMINAL 54 AMINO-ACID-RESIDUES BY ENDOGENOUS SERINE PROTEASE(S)

Bovine conglutinin, which is a serum lectin specific for N-acetylglucosamine, was purified from heat-inactivated bovine serum by Sepharose 4B-mannan affinity chromatography. Without prior heat inactivation, however, the same affinity chromatography yielded a modified conglutinin, which retained sugar-binding, but not conglutination activity. The modified lectin was designated conglutinin-N. The conglutinin-N subunit (38 kDa) was smaller than that of conglutinin (45 kDa). The apparent molecular mass was also reduced from 1000 (conglutinin) to 600 kDa (conglutinin-N). Conglutinin-N and conglutinin reacted equally with a rabbit anti-conglutinin antiserum. Comparison of the NH2-terminal sequences of conglutinin-N with the primary structure of conglutinin [Lee, Y.-M., Leiby, K.R., Allar, J., Paris, K., Lerch, B., and Okarma, T.B. (1991) J. Biol. Chem. 266, 2715-2723] revealed that the removal of the 54 NH2-terminal amino acids of conglutinin produced conglutinin-N. This conversion of conglutinin to conglutinin N was catalyzed by the eluate fraction of the first affinity chromatography from untreated bovine serum and was inhibited by serine protease inhibitors such as [4-aminophenyl]methanesulfonyl fluoride and C1 inhibitor. Conglutinin-N had neither conglutination activity nor binding activity to the sensitized erythrocyte-solid phase iC3b complex (EAiC3b), although it retained the binding activity to mannan in solution and the original binding specificity for N-acetylglucosamine. Cross-linking showed that conglutinin-N formed trimers of the 38-kDa subunits, whereas conglutinin formed higher multimers of 9-18 mer. These results indicated that deleting the NH2-terminal portion containing three cysteine residues by endogenous serine protease(s) produced some conformational changes in the entire molecule, which resulted in the loss of binding activity to EAiC3b and retention of the sugar-binding property. © 1993 Academic Press, Inc.