Probing the antibody-catalyzed water-oxidation pathway at atomic resolution.
Antibodies can catalyze the generation of hydrogen peroxide (H2O2) from singlet dioxygen (1O2*) and water via the postulated intermediacy of dihydrogen trioxide (H2O3) and other trioxygen species. Nine different crystal structures were determined to elucidate the chemical consequences to the antibody molecule itself of exposure to such reactive intermediates and to provide insights into the location on the antibody where these species could be generated. Herein, we report structural evidence for modifications of two specific antibody residues within the interfacial region of the variable and constant domains of different murine antibody antigen-binding fragments (Fabs) by reactive species generated during the antibody-catalyzed water oxidation process. Crystal structure analyses of murine Fabs 4C6 and 13G5 after UV-irradiation revealed complex oxidative modifications to tryptophan L163 and, in 4C6, hydroxylation of the Cgamma of glutamine H6. These discrete modifications of specific residues add further support for the "active site" of the water-oxidation pathway being located within the interfacial region of the constant and variable domains and highlight the general resistance of the antibody molecule to oxidation by reactive oxygen species generated during the water-oxidation process.
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Xueyong
X
Zhu
Paul
P
Wentworth
Anita D
AD
Wentworth
Albert
A
Eschenmoser
Richard A
RA
Lerner
Ian A
IA
Wilson
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000378:metabolism*; D002384:Catalysis; D018360:Crystallography, X-Ray; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008958:Models, Molecular; D010084:Oxidation-Reduction; D011487:Protein Conformation; D014867:Water / Q000378:metabolism*
D000906:Antibodies; D007140:Immunoglobulin Fab Fragments; D014867:Water