The activity of ribonuclease after digestion with carboxypeptidase

Abstract

On incubating ribonuclease (RNase) with a crystalline carboxypeptides preparation, a number of amino acids were liberated. Of the 17 amino acids in the RNase molecule, all except cystine and proline have thus far been quantitatively determined as products of this hydrolytic reaction with no loss in the original activity of the RNase. During the early phases of the digestion, the activity of the RNase was increased. A separation of the digested from the undigested RNase molecules has been achieved by chromatography of the digests in secondary-butyl alcohol: water (3:1) at 3° C for 15 hours. The native RNase molecules did not migrate under these conditions and were quite stable, whereas the digested RNase molecules migrated and were less stable. The data obtained with this procedure correlates reasonably well with the amino acid determinations as a measure of the per cent of the RNase molecules digested. The extent of digestion thus far achieved approximates 80% of the RNase molecules present, with no loss in enzymic activity. It is calculated that approximately 15% of the amino acid residues which constitute RNase are not essential for the enzymic activity, but appear to be necessary for the stability of the molecule.