Structural characterization of anti-inflammatory Immunoglobulin G Fc proteins

Creators: Ahmed, Alysia A.; Giddens, John; Pincetic, Andrew; Lomino, Joseph V.; Ravetch, Jeffrey V.; Wang, Lai-Xi; Bjorkman, Pamela J.

Abstract

Immunoglobulin G (IgG) is a central mediator of host defense due to its ability to recognize and eliminate pathogens. The recognition and effector responses are encoded on distinct regions of IgGs. The diversity of the antigen recognition Fab domains accounts for IgG's ability to bind with high specificity to essentially any antigen. Recent studies have indicated that the Fc effector domain also displays considerable heterogeneity, accounting for its complex effector functions of inflammation, modulation and immune suppression. Therapeutic anti-tumor antibodies, for example, require the pro-inflammatory properties of the IgG Fc to eliminate tumor cells, while the anti-inflammatory activity of Intravenous Immunoglobulin G (IVIG) requires specific Fc glycans for activity. In particular, the anti-inflammatory activity of IVIG is ascribed to a small population of IgGs in which the Asn297-linked complex N-glycans attached to each Fc C_H2 domain include terminal α2,6-linked sialic acids. We used chemoenzymatic glycoengineering to prepare fully di-sialylated IgG Fc and solved its crystal structure. Comparison of the structures of asialylated Fc, sialylated Fc, and F241A Fc, a mutant that displays increased glycan sialylation, suggests that increased conformational flexibility of the C_H2 domain is associated with the switch from pro- to anti-inflammatory activity of the Fc.

Additional Information

© 2014 Elsevier B.V. Received 25 April 2014, Revised 13 June 2014, Accepted 7 July 2014, Available online 15 July 2014. Edited by S. Sidhu. We thank Jost Vielmetter and the Caltech Protein Expression Center for protein production; Beth Stadtmueller and members of the Bjorkman, Ravetch, and Wang laboratories for critical reading of the manuscript; Marta Murphy for help making figures; and Jens Kaiser and members of the staff at the SSRL and the ALS for help with data collection and processing. Operations at SSRL are supported by the US Department of Energy and the National Institutes of Health. The ALS is supported by the Director, Office of Science, Office of Basic Energy Sciences, of the US Department of Energy under Contract No. DE-AC02-05CH11231. This research was supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health Grant HIVRAD P01 AI100148 (to P.J.B. and J.V.R.); (the content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health) the National Institutes of Health Grant R01 AI041239 (to P.J.B.), Award DP1OD006961 from the Office of the Director, National Institutes of Health (to P.J.B.), Grant R01 GM096973 (to L.X.W.), and Grant R56 AI034662 (to J.V.R.); and the Molecular Observatory at Caltech supported by the Gordon and Betty Moore Foundation. P.J.B. is a Howard Hughes Medical Institute investigator. Accession numbers: Atomic coordinates and structure factors have been deposited in the PDB with accession codes 4Q7D (wtFc), 4Q6Y (di-sFc), and 4Q74 (F241A Fc).

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