Racemization of the Succinimide Intermediate Formed in Proteins and Peptides: A Computational Study of the Mechanism Catalyzed by Dihydrogen Phosphate Ion

In proteins and peptides, D-aspartic acid (D-Asp) and D-beta-Asp residues can be spontaneously formed via racemization of the succinimide intermediate formed from L-Asp and L-asparagine (L-Asn) residues. These biologically uncommon amino acid residues are known to have relevance to aging and pathologies. Although nonenzymatic, the succinimide racemization will not occur without a catalyst at room or biological temperature. In the present study, we computationally investigated the mechanism of succinimide racemization catalyzed by dihydrogen phosphate ion, H2PO4-, by B3LYP/6-31+G(d,p) density functional theory calculations, using a model compound in which an aminosuccinyl (Asu) residue is capped with acetyl (Ace) and NCH3 (Nme) groups on the N- and C-termini, respectively (Ace-Asu-Nme). It was shown that an H2PO4- ion can catalyze the enolization of the H-alpha-C-alpha-C=O portion of the Asu residue by acting as a proton-transfer mediator. The resulting complex between the enol form and H2PO4- corresponds to a very flat intermediate region on the potential energy surface lying between the initial reactant complex and its mirror-image geometry. The calculated activation barrier (18.8 kcal.mol(-1) after corrections for the zero-point energy and the Gibbs energy of hydration) for the enolization was consistent with the experimental activation energies of Asp racemization.