Factors Influencing the Selection of α-Amino Acids over β-Amino Acids in Nature: Studies Involving Protein- l -isoaspartyl Methyltransferase Substrates and Related Peptides

To understand the differences resulting from the presence of an α- or β-amino acid in a peptide, we used two sets of amino acids corresponding to Asp/isoAsp and Ala/isoAla in two types of host peptides: one containing Gly (with no side chains) and the other containing Ala (representing amino acids with side chains), and measured their tendency to form fibrils. The peptides with a β-amino acid formed fibrils, while those with an α-amino acid did not exhibit this proclivity, as inferred from the thioflavin T (ThT) fluorescence intensity measurements. The fibrillation of the hexapeptide with isoAsp (the β-amino acid corresponding to Asp) was inhibited by protein-l-isoaspartyl methyltransferase (PIMT), a repair enzyme that converts the abnormal isoAsp residue to normal Asp. Isothermal Titration Calorimetry revealed the exothermic mode of binding of A6-isoAsp (Ala-based host hexapeptide) with PIMT. Far-UV CD spectroscopy revealed a β-sheet to α-helix transformation of isoAsp-containing peptides in the presence of PIMT. The hydrophobicity of the peptides was measured by noting their distribution in a mixture of water and octanol, where the β-amino acid exhibits a more positive hydrophobicity value than the α-amino acid. This may suggest that the hydrophobic forces bring chains containing β-amino acids together to enable nucleation for fibril formation. Molecular dynamics (MD) simulations indicated that isoAsp-based host peptides, formed into fibrils, have stable structures over the course of simulations, whereas the equivalent peptide with Asp disintegrated. Models of a single β-strand of these peptides bound to the active site of the enzyme were stable.