Biochemical characterization of a Ca(2+)-dependent lectin from the hemolymph of a photosymbiotic marine bivalve, Tridacna derasa (Röding).

A Ca(2+)-dependent lectin was purified from the hemolymph of a photosymbiotic bivalve, Tridacna derasa. An electrophoretically homogeneous form was obtained by using affinity chromatography with Sepharose 4B. More than 80% of the hemolymph protein was accounted for by this lectin. The apparent molecular mass of the lectin, in its native form exhibiting hemagglutinating activity, was estimated by gel filtration analysis to be approximately 480 kDa. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in the absence of reductants, it migrated as a single band corresponding to a very large size, while in the presence of 2-mercaptoethanol, it migrated as two distinct bands of 23 and 46 kDa. These results indicate that the subunits were linked by disulfide bridges to form the native protein. After reducing pyridylethylation, each of the 23- and 46-kDa polypeptides was isolated by gel filtration in a mobile phase containing guanidine-HCl. The two polypeptides had the same amino-terminal sequence and a similar amino-acid composition, and in the presence of 2-mercaptoethanol gave a single band on isoelectric focusing at a pH of 6.0. The results suggested that the 46-kDa peptide is a homodimer of 23-kDa subunits held together by a covalent bond other than a disulfide linkage. This lectin required calcium ions for its activity. By ultraviolet spectrophotometry the association constant for the calcium ion was determined to be 0.88 mM. The hemagglutinating activity decreased dramatically below pH 6.5, but re-increased to the original level when the solution was neutralized. Such a pH-dependent alteration of the ligand-binding activity was similar to that found in vertebrate asialoglycoprotein receptors.(ABSTRACT TRUNCATED AT 250 WORDS)