Alterations in the structure of the ribose moiety of ATP reduce its effectiveness as a substrate for the sarcoplasmic reticulum ATPase.

The substrate specificity of the calcium ATPase of isolated sarcoplasmic reticulum vesicles was examined using a series of ribose-modified ATP analogs. Steady state hydrolytic rate measurements using analogs modified at the 2'- and 3'-positions demonstrate that both hydroxyl groups contribute to substrate specificity and high catalytic activity. The 3'-hydroxyl is especially significant in this regard since all analogs modified at the 3'-position were slowly hydrolyzed, if at all, and they did not produce the activation at millimolar concentrations characteristically observed with ATP. In contrast, ATP analogs modified only at the 2'-position were more rapidly hydrolyzed (although at rates less than for ATP), and they did produce activation at millimolar concentrations. These results suggest that neither the catalytic (high affinity) nor the regulatory (low affinity) site of the CaATPase tolerates changes in the hydroxyl substituent at the 3'-position of ATP, whereas steady state rates associated with substrate binding at both types of sites are less affected by changes in the 2'-hydroxyl of ATP.