Watching Pseudomonas mevalonii HMG-CoA Reductase in Action

While static protein crystallography provides invaluable insights into the mechanism of enzyme catalyzed reactions, the bond forming and breaking steps are not observed directly, usually providing only indirect information on these steps.We present time-resolved crystallography studies of the thiohemiacetal formation in Pseudomonas mevalonii HMG-CoA Reductase at successive time points after initiation of the reaction using a pH jump trigger, demonstrating the formation of the thiohemiacetal carbon-sulfur bond in the crystal.Comparison of the structural data to MD simulations and theozyme calculations suggest that the reaction proceeds via a deprotonated CoA sulfur.The structure at 2 min 20 sec after initiation appears to be close to the activated complex where the concerted reactions of carbon-sulfur bond formation and proton transfer occur.Taken together, the time resolved crystallographic and computational data allow the direct observation of enzyme-catalyzed bond formation in a complex reaction mechanism in atomistic detail.