CARBONIC-ANHYDRASE FROM NICOTIANA-TABACUM LEAVES

In this study, carbonic anhydrase (CA: carbonate hydrolyase; E.C.4.2.1 .1) was purified from adult Nicotiana tabacum leaves and studied bioch emically. The enzyme was purified twice times by using (NH4)(2)SO4 pre cipation and DEAE-cellulose column chromatography, and its activity wa s determined for two different substrates (CO2 and p-nitrophenyl aceta te). The enzyme obtained from the ion exchange column was purified 40. 7 fold and the purity was controlled by 3%-10% discontinuous SDS-PAGE. The pH of the purified enzyme varied between 6.0 and 7.2, the optimum being 6.9. V-max and K-m values were calculated with p-nitrophenyl ac etate (0.1524 mM, 0.5446 mM, respectively) as substrate. The optimum t emperature for the enzyme was 40 degrees C. The molecular weight of th e enzyme and of the subunits were found to be approximately approximat e to 137.000 daltons and approximate to 22.800 daltons, respectively. The results indicate that 6 subunits are present. Changes in enzyme ac tivity were determined in the presence of caffeine, nicotine, metal io ns and some chemicals.