Crystal structure of a protein repair methyltransferase from Pyrococcus furiosus with its L-isoaspartyl peptide substrate.

Protein L-isoaspartyl (D-aspartyl) methyltransferases (EC 2.1.1.77) are found in almost all organisms. These enzymes catalyze the S-adenosylmethionine (AdoMet)-dependent methylation of isomerized and racemized aspartyl residues in age-damaged proteins as part of an essential protein repair process. Here, we report crystal structures of the repair methyltransferase at resolutions up to 1.2 A from the hyperthermophilic archaeon Pyrococcus furiosus. Refined structures include binary complexes with the active cofactor AdoMet, its reaction product S-adenosylhomocysteine (AdoHcy), and adenosine. The enzyme places the methyl-donating cofactor in a deep, electrostatically negative pocket that is shielded from solvent. Across the multiple crystal structures visualized, the presence or absence of the methyl group on the cofactor correlates with a significant conformational change in the enzyme in a loop bordering the active site, suggesting a role for motion in catalysis or cofactor exchange. We also report the structure of a ternary complex of the enzyme with adenosine and the methyl-accepting polypeptide substrate VYP(L-isoAsp)HA at 2.1 A. The substrate binds in a narrow active site cleft with three of its residues in an extended conformation, suggesting that damaged proteins may be locally denatured during the repair process in cells. Manual and computer-based docking studies on different isomers help explain how the enzyme uses steric effects to make the critical distinction between normal L-aspartyl and age-damaged L-isoaspartyl and D-aspartyl residues.