Analysis of a concerted mechanism in β-lactam enzymatic hydrolysis. A quantum mechanics/molecular mechanics study

One of the postulated mechanisms for the acylation step in β-lactamase catalyzed hydrolysis of β-lactams, a concerted one, has been explored by means of a quantum mechanics/molecular mechanics approach. Minima and transition structures for the reaction path are reported. The TEM-1 enzyme, a class A β-lactamase, and a penicillanate, a substrate easily hydrolyzed by this enzyme, constitute the system employed in our study. We have also analyzed the effects of the protonation state of Lys73 on the reaction mechanism. The energy barriers obtained, too high for a catalytic process, indicate that a concerted mechanism is not the most probable enzymatic mechanism for the acylation. Useful information is obtained by comparing the enzyme structures corresponding to the protonated and the deprotonated Lys73 residue along the reaction path. In the protonated Michaelis complex the Glu166 residue appears considerably closer to the Lys73 residue than in the deprotonated structure. This fact implies that an initially protonated Lys73 could easily transfer a proton and thus would not be a factor in excluding acylation mechanisms in which Lys73 acts as the general base in the deprotonation of Ser70. On the other hand, the Lys73 deprotonated acyl–enzyme structure is in better agreement with the reported X-ray crystallographic data than that of the protonated case.