The effects of aspartic acid‐bond isomerization on <i>in vitro</i> properties of the amyloid β‐peptide as modeled with <i>N</i>‐terminal decapeptide fragments

Abstract

The 42-amino acid A beta, the major constituent of the senile plaque deposits of the brains of Alzheimer's disease patients, exhibits a high degree of heterogeneity at its N-terminus. Isomerization of aspartic acid bonds at residues 1 and 7 renders A beta more prone to aggregate and form extended structure as it was shown by in vivo and in vitro studies. We recently demonstrated the ability of mid-chain aspartic acid-bond isomerization to break the dominant helical structure of the N-terminal decapeptide fragment by CD. In the current study we use molecular modeling to show that insertion of the extra -CH2-group into the decapeptide backbone results in the formation of stable reverse-turns and destabilizes the helical conformer that competes with the extended structure at the full-sized peptide level. The molecular modeling also reveals a limited propensity of the diisomerized peptide to form extended structure directly. Anti-A beta pAb 2332 is more sensitive for the non-isomerized status of the decapeptide than that of the full-sized peptide. mAb 6E10, raised against unmodified A beta recognizes only the unmodified decapeptide or the peptide isomerized at the first aspartic acid in a conformation-dependent manner, but does not recognize the mid-chain isomerized or diisomerized decapeptide in any circumstance. The diisomerized decapeptide was used as immunogen to generate polyclonal antibody 14943 that is not selective for the isomerized status of either the full-size peptide or the decapeptide, but recognizes the isomerized peptides preferentially when the peptide antigen structures are conserved during the enzyme-linked immunoassay procedure. Owing to the aberrant behavior of the full-sized A beta peptide during standard RP-HPLC, serum stability studies that indicate extracellular stability can be more effectively performed on the decapeptide fragments. Remarkably, the diisomerized peptide exhibits a significantly increased stability towards serum peptidases compared with the unmodified or monoisomerized peptides, suggesting a possible mechanism of the retention of the isomerized A beta peptide in the affected brains.