title: Crystal structure of a protein repair methyltransferase from Pyrococcus furiosus with its L-isoaspartyl peptide substrate.
abstract: 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.
authors.affiliation: Department of Chemistry and Biochemistry and Molecular Biology Institute, University of California, Los Angeles 90095-1569, USA.
authors.forename: S C
authors.initials: SC
authors.lastname: Griffith
authors.forename: M R
authors.initials: MR
authors.lastname: Sawaya
authors.forename: D R
authors.initials: DR
authors.lastname: Boutz
authors.forename: N
authors.initials: N
authors.lastname: Thapar
authors.forename: J E
authors.initials: JE
authors.lastname: Katz
authors.forename: S
authors.initials: S
authors.lastname: Clarke
authors.forename: T O
authors.initials: TO
authors.lastname: Yeates
mesh_terms: D000241:Adenosine / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D003067:Coenzymes / Q000378:metabolism; D018360:Crystallography, X-Ray; D026581:Isoaspartic Acid / Q000737:chemistry / Q000378:metabolism*; D007536:Isomerism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*; D010802:Phylogeny; D011487:Protein Conformation; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase / Q000737:chemistry* / Q000378:metabolism*; D020027:Pyrococcus furiosus / Q000201:enzymology*; D012435:S-Adenosylhomocysteine / Q000378:metabolism; D012436:S-Adenosylmethionine / Q000378:metabolism; D016415:Sequence Alignment; D012984:Software; D055672:Static Electricity; D013379:Substrate Specificity
chemical_list: D003067:Coenzymes; D026581:Isoaspartic Acid; D010455:Peptides; D012436:S-Adenosylmethionine; D012435:S-Adenosylhomocysteine; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase; D000241:Adenosine