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ENZYME ACTIVITY: A MOVING EXPERIENCE
Residue motions are found to be related to catalytic mechanism
STU BORMAN
The rapid motions of an enzyme's amino acid residues when it's in its catalytic transition state have been captured for the first time. Assistant professor of biochemistry Dorothee Kern of Brandeis University and co-workers used nuclear magnetic resonance to detect backbone motion in cyclophilin A and found movement in one residue to be strongly correlated with the catalytic mechanism [Science, 295, 1520 (2002)].
Researchers have previously studied static pictures (such as crystal structures) and motions of enzymes, but only in enzyme resting states, not during catalysis, Kern notes. "Because the transition state exists only fleetingly, enzyme dynamics during transition-state formation and decay have never been directly detected during an enzyme-catalyzed reaction in solution," explains Joseph J. Falke, professor of chemistry and biochemistry at the University of Colorado, in a Science commentary.
Kern and coworkers have now used NMR relaxation methods to look for motions in 160 amide nitrogens of the enzyme cyclophilin A during catalysis. They detected backbone movements in 10. In nine, these motions turned out to be associated primarily with substrate binding and unbinding. But rate analysis showed that movements of one residue, arginine-55, correlated strongly with catalysis. This makes sense, since arginine-55's side chain was already known to hydrogen-bond to the substrate and to play a key catalytic role.
The study analyzed backbone movements but not side-chain motions, which can also be directly involved in catalysis. But Kern and coworkers are currently extending the technique to side chains.
Protein motions have been a missing link in rational drug design, so pharmaceutical companies may be interested in the technique. As Kern explains, rational drug design has mainly been done by rigid-body docking, "but in the past 10 years, it's been shown that this usually doesn't work because you have to take into account the mobility of your protein and drug. This is the kind of problem we are trying to attack in the long term with our experiments--to really go beyond static pictures and explore the protein as a dynamic object."
MOTION PICTURE Motion of cyclophilin A's arginine-55 residue (black) correlates strongly with catalytic action: the cis-trans conversion of the enzyme's substrate (light and dark green). Other numbered residues (red) are among nine whose motions are associated with substrate binding and unbinding.
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