Biochemical characterization of purified carbonic anhydrase from the octocoral Leptogorgia virgulata

Carbonic anhydrase (CA) has been isolated and purified from the octocoral Leptogorgia virgulata (Lamarck) in an effort to investigate its role in the mineralization and demineralization of spicules and other calcified hard tissues. Affinity-chromatography using Prontosil-derivatized carboxymethylcellulose (CM) Bio-Gel A provided a one-step purification for 30 kdalton polypeptides with carbonic anhydrase activity. Four distinct polypeptides (designated , , , and ) are separated from one another at this molecular weight by two-dimensional gel electrophoresis. The specific activity of the L. virgulata CA is on average 57.5±1.5 U g^-1, is inhibitable by 10^-6 M acetazolamide, and is unaffected by 5mM dithiotheitol. The amino acid composition of these polypeptides is similar to that of mammal, bird, reptile, fish and arthropod species. Antiserum made against the L. virgulata CA reacts specifically with the 30 kdalton polypeptides in western blots, and crossreacts with human CA I and II. Antiserum against avian CA II crossreacts with the L. virgulata 30 kdalton polypeptides. This is the first report of the characterization of a purified CA from an octocoral, and production of a CA antiserum to a species in the phylum Cnidaria.