IG N-GLYCAN ORIENTATION CAN INFLUENCE INTERACTIONS WITH THE COMPLEMENT-SYSTEM

This study was prompted by the paradoxical observation that a pair of dinitrophenyl-specific murine monoclonal IgG2a Abs had similar monosac charide content and yet differed in their binding to lectins. The diff erential lectin-binding properties were lost when the Abs were denatur ed, suggesting that variations in lectin binding reflected the conform ational accessibility of the N-glycans rather than intrinsic differenc es in the lectin binding capacity of the glycans themselves. This hypo thesis was supported by experiments indicating that the degree to whic h the N-glycans on the Abs were reactive with beta-1,4-galactosyltrans ferase or susceptible to peptide N-glycosidase F corresponded directly to their relative accessibility to lectins, Moreover, the relative su sceptibility to these enzymes and accessibility to lectins was inverse ly related to the capacity of the Abs to activate the classical pathwa y, suggesting that the orientation of the more accessible N-glycan mig ht inhibit C1q binding, This hypothesis was supported by evidence that enzymatic cleavage of the more accessible N-glycan resulted in enhanc ed C1q, C4b, and C3b deposition. Conversely, removal of the less acces sible N-glycan expressed by the other Ab inhibited C1q, C4b, and C3b d eposition, The respective increase or decrease in C3b deposition on th e two deglycosylated Abs was magnified when complement activation was performed in factor B-depleted serum, suggesting that N-glycan conform ation primarily affects the classical pathway. Collectively, these dat a suggest that the orientation of the N-glycan expressed on Igs can pr ofoundly influence interaction with the complement system.