biostudies-literatureSubedi GPHHS | National Institutes of HealthRoy J. Carver Department of Biochemistry, Biophysics &amp; Molecular BiologyNIGMS NIH HHS16842-16850https://www.ebi.ac.uk/biostudies/studies/S-EPMC6204906biostudies-literatureUnknown293(43)Fc γ receptors (FcγRs) bind circulating IgG (IgG1) at the surface of leukocytes. Antibodies clustered at the surface of a targeted particle trigger a protective immune response through activating FcγRs. Three recent reports indicate that the composition of the asparagine-linked carbohydrate chains (<i>N</i>-glycans) of FcγRIIIa/CD16a impacted IgG1-binding affinity. Here we determined how <i>N</i>-glycan composition affected the affinity of the "low-affinity" FcγRs for six homogeneous IgG1 Fc <i>N</i>-glycoforms (G0, G0F, G2, G2F, A2G2, and A2G2F). Surprisingly, CD16a with oligomannose <i>N</i>-glycans bound to IgG1 Fc (A2G2) with a <i>K_D</i> = 1.0 ± 0.1 nm This affinity represents a 51-fold increase over the affinity measured for CD16a with complex-type <i>N</i>-glycans (51 ± 8 nm) and is comparable with the affinity of FcγRI/CD64, the sole "high-affinity" FcγR. CD16a <i>N</i>-glycan composition accounted for increases in binding affinity for the other IgG1 Fc glycoforms tested (10-50-fold). This remarkable sensitivity could only be eliminated by preventing glycosylation at Asn¹⁶² with an Asn-to-Gln mutation; mutations at the four other <i>N</i>-glycosylation sites preserved tighter binding in the Man5 glycoform. None of the other low-affinity FcγRs showed more than a 3.1-fold increase upon modifying the receptor <i>N</i>-glycan composition, including CD16b, which differs from CD16a by only four amino acid residues. This result indicates that CD16a is unique among the low-affinity FcγRs, and modifying only the glycan composition of both the IgG1 Fc ligand and receptor provides a 400-fold range in affinities.The Journal of biological chemistryCD16a with oligomannose-type <i>N</i>-glycans is the only "low-affinity" Fc γ receptor that binds the IgG crystallizable fragment with high affinity <i>in vitro</i>.PMC6204906R01 GM115489R01GM115489Start-up fundsBarb AWSubedi GPfalseCD16a with oligomannose-type <i>N</i>-glycans is the only "low-affinity" Fc γ receptor that binds the IgG crystallizable fragment with high affinity <i>in vitro</i>.Fc γ receptors (FcγRs) bind circulating IgG (IgG1) at the surface of leukocytes. Antibodies clustered at the surface of a targeted particle trigger a protective immune response through activating FcγRs. Three recent reports indicate that the composition of the asparagine-linked carbohydrate chains (<i>N</i>-glycans) of FcγRIIIa/CD16a impacted IgG1-binding affinity. Here we determined how <i>N</i>-glycan composition affected the affinity of the "low-affinity" FcγRs for six homogeneous IgG1 Fc <i>N</i>-glycoforms (G0, G0F, G2, G2F, A2G2, and A2G2F). Surprisingly, CD16a with oligomannose <i>N</i>-glycans bound to IgG1 Fc (A2G2) with a <i>K_D</i> = 1.0 ± 0.1 nm This affinity represents a 51-fold increase over the affinity measured for CD16a with complex-type <i>N</i>-glycans (51 ± 8 nm) and is comparable with the affinity of FcγRI/CD64, the sole "high-affinity" FcγR. CD16a <i>N</i>-glycan composition accounted for increases in binding affinity for the other IgG1 Fc glycoforms tested (10-50-fold). This remarkable sensitivity could only be eliminated by preventing glycosylation at Asn¹⁶² with an Asn-to-Gln mutation; mutations at the four other <i>N</i>-glycosylation sites preserved tighter binding in the Man5 glycoform. None of the other low-affinity FcγRs showed more than a 3.1-fold increase upon modifying the receptor <i>N</i>-glycan composition, including CD16b, which differs from CD16a by only four amino acid residues. This result indicates that CD16a is unique among the low-affinity FcγRs, and modifying only the glycan composition of both the IgG1 Fc ligand and receptor provides a 400-fold range in affinities.2018-01-01T00:00:00Z2018 Oct2026-05-06T00:17:35.62Z2019-11-05T08:04:18ZS-EPMC62049063021386210.1074/jbc.ra118.00499810.1074/jbc.RA118.004998