Abstract (EN):
Human angiotensin-converting enzyme (ACE) is a zinc metallopeptidase that converts angiotensin I to the vasoconstrictor angiotensin II and inactivates the vasodilator bradykinin. This dual ability is vital to blood pressure regulation and management of hypertension. Despite the many enzymatic studies on zinc metallopeptidases, the correct substrate binding mode and catalysis of ACE are still not completely understood. Two buried chloride ions activate the ACE hydrolysis efficiency in a substrate-dependent manner, but the molecular mechanism associated with this activation also remains unclear. In this work, the catalytic mechanism of ACE was studied with atomistic detail, using a hybrid quantum mechanical/molecular mechanical method at the ONIOM(M06-2X/6-311+G(d,p):Amber//B3LYP/6-31G(d):Amber) level. The hydrolytic reaction proceeds via a general acid/base mechanism, in which the first mechanistic step involves the displacement of the zinc-bound water molecule that performs a nucleophilic attack on the scissile carbonyl bond to form an oxyanion that results in a gem-diol intermediate. The second step involves a proton transfer from Glu384 to the peptide nitrogen and a subsequent cleavage of the peptidic bond to yield the products in their neutral forms. The conserved residue Glu384 is ideally aligned and has the ability to slightly rearrange its conformation to act as a highly effective proton shuttle. Our results indicate that the nucleophilic attack is the rate-limiting step of ACE catalysis (barrier of approximate to 19 kcal/mol), which agrees with the experimental data available. Molecular dynamics simulations on ACE were also performed, and the data reported here provide a structural basis for the chloride-dependent activity of ACE. It was observed that the Cl2 absence allows a conformational rearrangement of the Arg522 side chain, which subsequently makes an electrostatic interaction with the zinc-bound Glu411 and perturbs the metal center polarization role during catalysis.
Language:
English
Type (Professor's evaluation):
Scientific
No. of pages:
11