Role of oligosaccharide residues of IgG1-Fc in Fc gamma RIIb binding

Engagement of Fc gamma receptors (Fc gamma Rs) with the Fe region of IgG el icits immune responses by leukocytes. The recent crystal structure of Fc ga mma RIII in complex with IgG-Fc has provided details of molecular interacti ons between these components (Sondermann, P., Huber, R., Oosthuizen, V., an d Jacob, U. (2000) Nature 406, 267-273). One of the most intriguing issues is that glycosylation of IgG-Fc is essential for the recognition by Fc gamm a Rs although the carbohydrate moieties are on the periphery of the Fc gamm a RIII-Fc interface. To better understand the role of Fe glycosylation in F c gammaR binding we prepared homogeneous glycoforms of IgG-Fc (Cri) and inv estigated the interactions with a soluble form of Fc gamma RIIb (sFc gamma RIIb). A 1:1 complex stoichiometry was observed in solution at 30 degreesC (K-d, 0.94 muM; DeltaG, - 8.4 kcal mol(-1); DeltaH, -6.5 kcal mol(-1); T De ltaS, 1.9 kcal mol(-1); DeltaC(p), -160 cal mol(-1) K-1). Removal of termin al galactose residues did not alter the thermodynamic parameters significan tly. Outer-arm GlcNAc residues contributed significantly to thermal stabili ty of the C(H)2 domains but only slightly to sFc gamma RIIb binding. Trunca tion of 1,3- and 1,6-arm mannose residues generates a linear trisaccharide core structure and resulted in a significantly decreased affinity, a less e xothermic DeltaH, and a more negative DeltaC(p) for sFc gamma RIIb binding, which may result from a conformational change coupled to complex formation . Deglycosylation of the C(H)2 domains abrogated sFc gamma RIIb binding and resulted in the lowest thermal stability accompanied with noncooperative u nfolding. These results suggest that truncation of the oligosaccharides of IgG-Fc causes disorder and a closed disposition of the two C(H)2 domains, i mpairing sFc gamma RIIb binding.