Activation of bovine muscle carbonic anhydrase by modification of thiol groups

Two of the five cysteine residues in bovine muscle carbonic anhydrase (isoenzyme III) react rapidly and stoichiometrically with Ellman's reagent without effect on the CO2 hydration activity. These residues, which can be alkylated with iodoacetamide, have been identified as Cys-183 and Cys-188. Treatment of the enzyme with a large excess of Ellman's reagent results in additional derivatization of thiol groups and a 180% increase of the CO2 hydration activity. The cysteine residues associated with this activation are Cys-66 as well as Cys-203 and/or Cys-206. Activation has also been achieved with 2,2'-dithiodipyridine (120%) and with methyl methanethiosulfonate (approx. 400%), whereas no activation could be obtained with iodoacetamide, iodoacetate or N-ethylmaleimide.