Exploring Functions for Glycosylation in Host Defence
325
5
The Structure and Function of the Fc Oligosaccharides in IgA1 and IgG
is Influenced by the Location of the Glycans
(i) The sugars attached to IgAI CH2 domains are larger than those on IgG
and, in contrast to those on IgG, are not contained within the CH2 domains
Crystallographic studies of human IgG Fc (Deisenhofer, 1981) have shown that
the two CH2 domains do not form extensive lateral associations. The resulting
interstitial region accommodates the oligo saccharides, which are attached to
Asn297 on each heavy chain (Fig. 23.3a). In contrast, in IgAI (Fig. 23.3b), the
interstitial space is not large enough to accommodate the sugars. Moreover,
molecular modelling (Mattu, et aI., 1998) indicates that the Asn298 side chains
point away from the protein surface such that the N -glycans are fully exposed on
the outside of the molecule. As a consequence, IgA1 sugars can be larger than
those on IgG (Fig 23.4) (Field et aI., 1994, Mattu et al., 1998).
(ii) IgG Fc Oligo saccharides Maintain the Relative Geometry of the CH2
Domains.
In contrast to IgA1, in IgG Fc the oligo saccharides maintain the relative geometry
of the CH2 domains. The oligo saccharides in the Fc region of IgG are of the complex bi-antennary type and can be classified according to the number of terminal
galactose residues they contain. Each sugar has either two terminal galactose residues (G2), one galactose and one GlcNAc (Gl) or two terminal GlcNAc residues
(GO). The major determinant for the binding site for the Fc receptor is Leu235
and the receptor may interact directly with the region linking the CH2 domain to
the hinge (Duncan et al., 1988; Burton and Woof, 1992). Both non-glycosylated
and degalactosylated IgG bind less efficiently to the Fcy receptors (Nose and Wigzell1983, Leatherbarrow et aI., 1984, Tsuchiya et aI., 1989). This is consistent with
the finding that protein-oligosaccharide interactions involving the conserved glycans at Asn297 (Padlan, 1994) playa role in maintaining the geometry of the
hinge region and therefore of the CH2 domains in IgG Fc (Rudd et aI., 1991). In
contrast to IgG, in IgA1 no reduction in binding to its Fc receptor was detected
in CHO-K1 mutants in which the N-linked sugar sites in the CH2 domains had
been deleted (Mattu et aI., 1998). This result is not unexpected since the sugars in
the CH2 domains do not playa role in maintaining the hinge structure in IgAl.
(iii) Sugars Protect IgG and IgAI Against Proteolysis
Although the N-glycans in the CH2 domains of IgA1 do not playa role in maintaining domain structure, just as the multiple O-glycans in the hinge have been
shown to protect the hinge region of IgA1 against non-specific proteolysis (Mestecky and Kilian, 1985) so the N -glycans are expected to shield large areas of the
Fc region from proteases.
In the case of IgG, which can be cleaved by papain into Fab (the antigen binding fragment) and Fc (the region responsible for effector functions of the molecule, such as Fcy receptor binding and complement activation), non-glycosylated
IgG, expressed in tunicamycin treated cells, is more readily cleaved by papain
than normal IgG (Leatherbarrow and Dwek 1984). This indicates that the conserved glycans in the Fc provide protection against proteases.
325
5
The Structure and Function of the Fc Oligosaccharides in IgA1 and IgG
is Influenced by the Location of the Glycans
(i) The sugars attached to IgAI CH2 domains are larger than those on IgG
and, in contrast to those on IgG, are not contained within the CH2 domains
Crystallographic studies of human IgG Fc (Deisenhofer, 1981) have shown that
the two CH2 domains do not form extensive lateral associations. The resulting
interstitial region accommodates the oligo saccharides, which are attached to
Asn297 on each heavy chain (Fig. 23.3a). In contrast, in IgAI (Fig. 23.3b), the
interstitial space is not large enough to accommodate the sugars. Moreover,
molecular modelling (Mattu, et aI., 1998) indicates that the Asn298 side chains
point away from the protein surface such that the N -glycans are fully exposed on
the outside of the molecule. As a consequence, IgA1 sugars can be larger than
those on IgG (Fig 23.4) (Field et aI., 1994, Mattu et al., 1998).
(ii) IgG Fc Oligo saccharides Maintain the Relative Geometry of the CH2
Domains.
In contrast to IgA1, in IgG Fc the oligo saccharides maintain the relative geometry
of the CH2 domains. The oligo saccharides in the Fc region of IgG are of the complex bi-antennary type and can be classified according to the number of terminal
galactose residues they contain. Each sugar has either two terminal galactose residues (G2), one galactose and one GlcNAc (Gl) or two terminal GlcNAc residues
(GO). The major determinant for the binding site for the Fc receptor is Leu235
and the receptor may interact directly with the region linking the CH2 domain to
the hinge (Duncan et al., 1988; Burton and Woof, 1992). Both non-glycosylated
and degalactosylated IgG bind less efficiently to the Fcy receptors (Nose and Wigzell1983, Leatherbarrow et aI., 1984, Tsuchiya et aI., 1989). This is consistent with
the finding that protein-oligosaccharide interactions involving the conserved glycans at Asn297 (Padlan, 1994) playa role in maintaining the geometry of the
hinge region and therefore of the CH2 domains in IgG Fc (Rudd et aI., 1991). In
contrast to IgG, in IgA1 no reduction in binding to its Fc receptor was detected
in CHO-K1 mutants in which the N-linked sugar sites in the CH2 domains had
been deleted (Mattu et aI., 1998). This result is not unexpected since the sugars in
the CH2 domains do not playa role in maintaining the hinge structure in IgAl.
(iii) Sugars Protect IgG and IgAI Against Proteolysis
Although the N-glycans in the CH2 domains of IgA1 do not playa role in maintaining domain structure, just as the multiple O-glycans in the hinge have been
shown to protect the hinge region of IgA1 against non-specific proteolysis (Mestecky and Kilian, 1985) so the N -glycans are expected to shield large areas of the
Fc region from proteases.
In the case of IgG, which can be cleaved by papain into Fab (the antigen binding fragment) and Fc (the region responsible for effector functions of the molecule, such as Fcy receptor binding and complement activation), non-glycosylated
IgG, expressed in tunicamycin treated cells, is more readily cleaved by papain
than normal IgG (Leatherbarrow and Dwek 1984). This indicates that the conserved glycans in the Fc provide protection against proteases.