The ability of the human body to fight infection must be precisely regulated to be sensitive to pathogens but avoid spurious activation, tissue damage and disease. This balance is mediated, in large part, at the point of immunoglobulin G (IgG) recognition by the pro-inflammatory Fc? receptors (Fc?Rs), though fundamental questions about this association exist. Blocking the Fc-Fc?R interaction promises to offer new treatments for autoimmune disorders, however, a description of the factors contributing to Fc binding by Fc?R, namely the role of the essential Fc Asn-297 N-glycan, has not been described. This study aims to elucidate the role of the N-glycan in binding, and by extension, immune system activation. This will be of high impact and critical to understanding the mechanism used by the body to regulate immune system activation. The first step in this project is to develop a method to express structural-biology quantities of 15N-enriched, natively N-glycosylated IgG1 Fc fragment. Following purification, the IgG N-glycans will be remodeled enzymatically to homogeneity using UDP-13C-galactose. Next, we will assess the affect of Fc?RIII on the conformations of the IgG1 Fc N- glycan using solution nuclear magnetic resonance spectroscopy measurements of glycan motion and structure. Fc?RIII will be prepared and titrated into the IgG1 Fc fragment containing 13C-galactose terminated N-glycans. Finally, we will characterize the role of the Fc?RIII extracellular domain binding on the structure of the total IgG1 Fc glycoprotein. Structural and motional alterations in the polypeptide will be measured using the 15N probes incorporated into the polypeptide. These data will directly impact human health and lead to new approaches to modulate the Fc-Fc?R interaction and thus autoimmune diseases, as well as tuning of therapeutic antibodies to have desirable immunoregulatory properties. Current research focuses on describing the structure and motion of the IgG1 Fc N-glycan on the Fc only and the Fc in a complex with a Fc-binding peptide. Autoimmune disorders result from improper immune system activation and are debilitating diseases that limit the health and productivity of affected individuals. This proposal will utilize recent discoveries to study the molecular details of immune system activation and will guide the future development of more effective treatments for these disorders.

Public Health Relevance

Autoimmune disorders result from improper immune system activation and are debilitating diseases that limit the health and productivity of affected individuals. This proposal will utilize recent discoveries to study the molecular details of immune system activation and will guide the future development of more effective treatments for these disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Career Transition Award (K22)
Project #
1K22AI099165-01
Application #
8280530
Study Section
Allergy & Clinical Immunology-1 (AITC)
Program Officer
Prograis, Lawrence J
Project Start
2013-01-18
Project End
2014-12-31
Budget Start
2013-01-18
Budget End
2013-12-31
Support Year
1
Fiscal Year
2013
Total Cost
$159,347
Indirect Cost
$9,347
Name
Iowa State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
005309844
City
Ames
State
IA
Country
United States
Zip Code
50011
Frank, Martin; Walker, Ross C; Lanzilotta, William N et al. (2014) Immunoglobulin G1 Fc domain motions: implications for Fc engineering. J Mol Biol 426:1799-811
Subedi, Ganesh P; Hanson, Quinlin M; Barb, Adam W (2014) Restricted motion of the conserved immunoglobulin G1 N-glycan is essential for efficient Fc?RIIIa binding. Structure 22:1478-88