Mapping of the binding site for FcµR in human IgM-Fc.

ABSTRACT

FcµR is a high-affinity receptor for the Fc portion of human IgM. It participates in B cell activation, cell survival and proliferation, but the full range of its functions remains to be elucidated. The receptor has an extracellular immunoglobulin (Ig)-like domain homologous to those in Fcα/µR and pIgR, but unlike these two other IgM receptors which also bind IgA, FcµR exhibits a binding specificity for only IgM-Fc. Previous studies have suggested that the IgM/FcµR interaction mainly involves the Cµ4 domains with possible contributions from either Cµ3 or Cµ2. To define the binding site more precisely, we generated three recombinant IgM-Fc proteins with specific mutations in the Cµ3 and Cµ4 domains, as well as a construct lacking the Cµ2 domains, and analyzed their interaction with the extracellular Ig-like domain of FcµR using surface plasmon resonance analysis. There is a binding site for FcµR in each IgM heavy chain. Neither the absence of the Cµ2 domains nor the quadruple mutant D340S/Q341G/D342S/T343S (in Cµ3 adjacent to Cµ2) affected FcµR binding, whereas double mutant K361D/D416R (in Cµ3 at the Cµ4 interface) substantially decreased binding, and a single mutation Q510R (in Cµ4) completely abolished FcµR binding. We conclude that glutamine at position 510 in Cµ4 is critical for IgM binding to FcµR. This will facilitate discrimination between the distinct effects of FcµR interactions with soluble IgM and with the IgM BCR.