Collect. Czech. Chem. Commun. 2003, 68, 2055-2079

Theoretical Studies of Catalysis by Carboxypeptidase A: Could Gas-Phase Calculations Support a Mechanism?

Alexandra Kilshtain-Vardia,b, Gil Shohama and Amiram Goldblumb,*

a Department of Inorganic and Analytical Chemistry, Institute of Chemistry, Hebrew University of Jerusalem, Israel 91120
b Department of Medicinal Chemistry and the David R. Bloom Center for Pharmacy, School of Pharmacy, Hebrew University of Jerusalem, Israel 91120


We compare recent quantum mechanical computations of alternative reaction pathways for carboxypeptidase A, a zinc proteinase, in an "enzyme environment" to similar calculations in the "gas phase" that include the minimal chemical entities that are required for a non-catalytic reaction. The main question that we address is whether anything may be learned from such reduced representations. Two general acid-general base alternative pathways and one nucleophilic pathway are compared. The original calculations were run on a relatively large model (120 atoms) of the active site of carboxypeptidase A which included zinc and its ligands, as well as the residues Arg145, Arg127, Glu270, a water molecule and a model dipeptide. The "gas-phase" pathways include only the dipeptide, water and Glu270 and serve as models for the non-catalytic pathway. The calculations were performed by semiempirical MNDO/H/d that includes modifications for d-orbital representations as well as for intra- and intermolecular multiple H-bond formation. The gas-phase results strengthen our previous conclusion about the preference for general acid-general base pathways for peptide cleavage by carboxypeptidase A rather than a "direct nucleophilic" pathway. The bottleneck of the reaction is proton transfer to the nitrogen in the peptide bond, preceding the peptide cleavage.

Keywords: Carboxypeptidase A; Enzymes; Proteinases; Inhibitors; Catalytic mechanism; MNDO/H; MNDO/d; General acid-general base; Reaction coordinate; Gas phase; Semiempirical calculations; Peptidomimetics.

References: 62 live references.