Calcium binding constituents of the organic shell matrix from the freshwater snail Biomphalaria glabrata

Abstract

The Ca 2+ binding of an EDTA-free water-soluble (SM) and -insoluble (IM) organic matrix of the freshwater snail Biomphalaria glabrata was investigated, using a 45Ca 2+ autoradiography after SDS-electrophoretical separation and a calcium binding assay. Electrophoresis of the SM showed a considerable amount of Alcian blue and Stains all positive material, regarded as glycosaminoglycans (GAGs) or proteoglycans (PGs). This part of the SM was slightly positive after 45Ca 2+ autoradiography at pH 6.8. The Ca 2+ binding increased, raising the pH to 7.4 and 8.0 and was especially strong when simulating the real conditions of the extrapallial space with a carbonate buffer of pH 7.4. The Ca 2+ binding assay of the IM showed the same pH-dependency that was observed in the SM. The titration of the IM with Ca 2+ at pH 8.0 lead to a dissociation constant of 7.5×10 −5 M. While Mg 2+ displaced 45Ca 2+ in the same way as nonradioactive Ca 2+, an ≈400-fold amount of Na + was necessary to reduce the binding of 45Ca 2+ to 50%. The Ca 2+ binding of the organic matrix from the B. glabrata shell appears to be a process of low specificity, medium affinity and high pH-dependency. Apparently, acidic carbohydrate-rich PGs are the only calcium binding constituents of the organic shell matrix.