Michael Harris, Ph.D.

Professor in Chemistry
University of Florida


Title: Altered (transition) States: Comparison of ribozyme and ribonuclease catalysis at the level of transition state structure reveals unique features relative to non-enzymatic catalysis


In many cases enzymes that catalyze the same chemical reaction can have very different active sites.  A prime example are protein ribonucleases and their catalytic RNA cousins (ribozymes).  Both protein ribonucleases (like RNase A) and small self-cleaving ribozymes (such as the HDV ribozyme) catalyze RNA 2’-O-transphosphorylation in which the RNA 2’-hydroxyl group attacks the adjacent phosphoryl and displaces the 5’ oxygen resulting in strand cleavage.  Both RNases and ribozymes are proposed to use similar catalytic modes (acid/base catalysis, metal ion catalysis, reducing conformational entropy, etc.) but appear to implement them in different ways, raising the fundamental question of whether they stabilize the same or different active sites.  To address this question we have used kinetic isotope effects to benchmark QM/MM simulations aimed at determining the transition states for non-enzymatic, ribonuclease catalyzed and ribozyme catalyzed RNA strand cleavage.  This work resulted in the first experimentally established transition state for the well-studied enzyme RNase A, and showed that ribozymes can stabilize a transition state that is very different from its protein counterparts.  The new information not only challenges our assumptions regarding the diversity of chemical transition states employed by enzymes, but also helps shed light on design principles for developing artificial catalysts that mimic the power of enzymes.