HORNE, F. R.; Texas State University: Localization of carbonic anhydrase in the crustacean cuticle

An organic matrix of protein and polysaccharides along with minerals is secreted extracellularly by decapod hypodermal cells. These macromolecules are the sites of ion concentration, crystal nucleation, crystal growth, and control of crystal morphology. Crustaceans manufacture and calcify a new shell in a matter of several days and the exoskeleton often consists of more than 40% organic matrix with the balance of the shell being calcium carbonate. Calcium ions are pumped from the environment into body fluids and then out again at the site of mineral deposition via the hypodermis. Bicarbonate secreted to maintain electrical neutrality is the source of carbonate ions. The calcium and carbonate ion product must exceed its solubility product to sustain crystal growth. Two issues arise in association with carbonate ions: a) there must be an adequate supply of bicarbonate ions to provide carbonate ions, and b) there must be a means to remove the proton as carbonate is formed from bicarbonate. Without the enzyme carbonic anhydrase (CA) the dehydration and hydration of carbon dioxide is a rather slow reaction. An extracellular CA secreted along with the matrix proteins might solve this dilemma. In this study cuticles were decalcified with formic acid, sectioned and treated with antibodies to bovine RBC CA II. Using CA antibodies the enzyme appeared to be localized through out the cuticle matrix of american lobster, Homarus americanus, blue crab, Callinectes sapidus, and crayfish, Procambarus clarkii. Non-calcified tergum and controls did react with antibodies and did not seem to have CA. Extracellular CA may function along with intracellular CA in maintaining bicarbonate levels and/or removing protons. CA might be especially important in fresh water with pH near 7.