Probing the antibody-catalyzed water-oxidation pathway at atomic resolution
- Xueyong Zhu†,
- Paul Wentworth, Jr.‡,§,
- Anita D. Wentworth‡,
- Albert Eschenmoser†,¶,∥,
- Richard A. Lerner†,‡,§,¶, and
- Ian A. Wilson†,§,¶
- Departments of †Molecular Biology and ‡Chemistry and ¶The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037; and ∥Laboratorium für Organische Chemie, Eidgenössische Technische Hochschule Hönggerberg HCl-H309, Universitätstrasse 16, CH-8093 Zurich, Switzerland
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Contributed by Richard A. Lerner, December 19, 2003
Abstract
Antibodies can catalyze the generation of hydrogen peroxide (H2O2) from singlet dioxygen (1O*2) 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 Cγ 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.
Footnotes
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↵ § To whom correspondence may be addressed. E-mail: wilson{at}scripps.edu, paulw{at}scripps.edu, or foleyral{at}scripps.edu.
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Abbreviations: Fab, antibody antigen-binding fragment; PEG, polyethylene glycol; SSRL, Stanford Synchrotron Radiation Laboratory.
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Data deposition: The six structures and structure factors of 4C6 Fab and two structures and structure factors of 13G5 Fab have been deposited at the Protein Data Bank, www.rcsb.org (PDB ID codes 1ru9, 1rua, 1ruk, 1rul, 1rum, 1rup, 1ruq, and 1rur, respectively).
- Copyright © 2004, The National Academy of Sciences
