Isolation and properties of a mannan-binding lectin from the coelomic fluid of the holothurian Cucumaria japonica

A new 44-kD, C-type mannan-binding lectin (MBL-C) consisting of two identic al subunits was isolated from the coelomic fluid of the holothurian Cucumar ia japonica. In the direct hemagglutination assay, the lectin was effective ly inhibited by highly branched mannans similar in structure to yeast manna ns and composed of alpha-(1-->2)- and alpha-(1-->6)bound D-mannopyranose re sidues. Hemagglutination was not inhibited by mannosaccharides, common cons tituents of the hydrocarbon chains of "normal" glycoproteins. The lectin re action depends on Ca2+ concentration: maximum activity of MBL-C is observed at 10 mM Ca2+. The activity of MBL-C increases in the pH range from 5 to 7 and reaches maximum at pH 7.0. The lectin is sensitive to temperature. Hea ting of the lectin solution at temperatures above 40 degrees C decreases ac tivity, while incubation at 90 degrees C for 1 h leads to complete irrevers ible inactivation. Carbohydrate specificity, Ca2+-dependence, and amino aci d composition indicate that MBL-C belongs to the C-type mannan-binding lect ins. Polyclonal antibodies against MBL-C revealed its immunochemical simila rity to a mannan-binding lectin from another holothurian species, Stichopus japonicus; this provides evidence for structural homology between these pr oteins.