Single molecule Förster resonance energy transfer studies of the effect of EndoS deglycosylation on the structure of IgG.

The bacterial enzyme EndoS specifically cleaves glycans bound to immunoglobulin G (IgG) molecules. Because this deglycosylation procedure leads to a diminished immune response, this enzyme has potential applications as a therapeutic for autoimmune disorders. Although the diminished immune response is attributed to a structural change in the Fc region of IgG antibodies, the specific nature of this structural change is not known due to the variety of results obtained by different experimental approaches. In order to better understand how EndoS deglycosylation impacts the structure of the Fc region of IgG antibodies, we have conducted single molecule Förster resonance energy transfer (FRET) studies of dye-labeled, freely diffusing antibodies. A comparison of the FRET efficiency histograms obtained for glycosylated and EndoS deglycosylated antibodies indicates that the Fc region can take on a wider variety of structures upon deglycosylation. This is demonstrated by the presence of additional peaks in the FRET efficiency histogram for the deglycosylated case.