HORNE, F. R.; Texas State University: Carbonic anhydrase in slipper lobster cuticles

The organic matrix of slipper lobsters, Scyllarides latus , serves as a site of ion concentration, crystal nucleation, crystal growth, and control of crystal morphology. Following molt crustaceans like the slipper lobster manufacture and calcify a new cuticle in a matter of days. Exoskeletal composition may consist of 40% organic matrix with the balance primarily of calcium carbonate. Calcium and bicarbonate ions must be mobilized at the site of mineral deposition for mineralization to happen in a matter of hours. Since the calcium and carbonate ion product must exceed its solubility product to sustain crystal growth, a) there must be an adequate supply of bicarbonate ions to provide carbonate ions, and b) there must be a means to remove protons. Carbonic anhydrase (CA) satisfies both of these requisites. Non-enzymatic dehydration and hydration of carbon dioxide is a rather slow reaction, so an extracellular CA secreted along with matrix proteins might solve this dilemma. In this study cuticles (intermolt cuticle, new uncalcified cuticle, and old decalcified cuticle) were sectioned following standard histological techniques and treated with antibodies to bovine RBC CA II. Use of CA antibodies to localize the cuticle enzyme showed that only the highly calcified intermolt cuticles gave a strong positive reaction to the CA antibody. Neither the new forming cuticle or the old cuticle appeared to respond to the CA antibodies. Apparently CA is secreted following molt during mineralization since the new unmineralized cuticle does not stain for CA. Mineral and nutrient reabsorption from the old cuticle prior to molt appears to remove any positive reaction to CA. These data along with those showing weak reactions to CA in poorly mineralized cuticles suggest that extracellular CA may function along with intracellular CA in maintaining bicarbonate levels and/or removing protons.