Side-chain conformers to allow conversion from normal to isoaspartate in age-related proteins and peptides

Dβ (or D-iso)- and Lβ- (or iso)- aspartyl (Asp) residues are accumulated in aged lens crystallins and amyloid beta (Aβ) proteins, respectively, as a result of spontaneous, nonenzymatic isomerization of normal Lα-Asp. To explore why such uncommon Asp isomers are accumulated, the stability of Lα-, Lβ-, and Dβ-Asp was compared in view of the staggered side-chain conformers. By using cylindrin (KVKVLGD⁷VIEV) from αB-crystallin and Aβ17-25 (L¹⁷VFF²⁰AED²³)VG²⁵) containing Asp isomers, the vicinal spin-spin coupling constants of Asp H_α-H_β1 and H_α-H_β2 were quantified by high-resolution solution ¹H NMR. It was found that the trans conformer was extremely preferred in Dβ-Asp7 side-chain of cylindrin. In Aβ17-25, the side chain of Lβ-Asp23 was likely to adopt trans conformer, while gauche conformers were rather rich in Lα-Asp23. In gauche conformers, the close distance between Asp carboxylate carbon (C_COO-) and backbone nitrogen (N) next to Asp is advantageous to the intramolecular cyclization to form succinimide intermediate, followed by the conversion from α- to β-Asp. The cyclization is limited in the trans conformer because of the long distance between C_COO- and N, to keep Dβ- or Lβ-Asp stable. This would be the reason for the site specificity of Asp isomerization in proteins. The higher population of trans conformer in Asp side chain, the less isomerization of Asp as shown as Asp76 in αA-crystallin. The stability and less reactivity of normal Asp and its isomers are the potential factors to determine whether or not the abnormal accumulation is permitted in aged crystallins and Aβ.