PURIFICATION AND PROPERTIES OF DOUBLE-STRANDED RNA-SPECIFIC ADENOSINE-DEAMINASE FROM CALF THYMUS

A double-stranded RNA-specific adenosine deaminase, which converts adenosine to inosine, has been purified to homogeneity from calf thymus. The enzyme was purified approximate to 340,000-fold by a series of column chromatography steps. The enzyme consists of a single polypeptide with a molecular mass of 116 kDa as determined by electrophoresis on a SDS/polyacrylamide gel. The native protein sediments at 4.2 s in glycerol gradients and has a Stokes radius of 42 Angstrom upon gel-filtration chromatography. This leads to an estimate of approximate to 74,100 for the native molecular weight, suggesting that the enzyme exists as a monomer in solution. Enzyme activity is optimal at 0.1 M KCl and 37 degrees C. Divalent metal ions or ATP is not required for activity. The K-m for double-stranded RNA substrate is approximate to 7 x 10(-11) M. The V-max is approximate to 10(-9) mol of inosine produced per min per mg and the K-cat is 0.13 min(-1).