Expression of Cytosolic and Non-Cytosolic Carbonic Anhydrase Enzymes from Bovine Large Intestine

Abstract

Carbonic anhydrase is an enzyme that takes responsibility in inhalation function but, until today, carbonic anhydrase is not examined if it is present in the bovine large intestine or not. The enzyme carbonic anhydrase was purified and separately characterized according to the bonding forms in 4 steps such as outer peripheral, cytosolic, inner peripheral and integral. Affinity chromatography was used for purification of the enzyme in all four steps. The affinity column was prepared with sepharose-4B-L-tyrosine-sulphanilamide. Purified carbonic anhydrase was obtained at each step. Enzyme activity was measured by CO2 hydratase activity and esterase activity methods. Optimum pH and optimum temperature were defined for purified enzymes at each step. Morover molecular weight and purity were detected by gel filtration and SDS-PAGE electrophorose. In addition, the enzyme's Km and Vmax values were found with the Lineweaver-Burk method. The results are discussed in comparison with other mammalian carbonic anhydrases. Carbonic anhydrase was shown to be exist in bovine large intestine and this enzyme was optimized.