Expression of biological effector functions by immunoglobulin G molecules lacking the hinge region.

Abstract

Several biological effector functions mediated by sites on the Fc region of human IgG1 have been studied in two variant IgGlK monoclonal proteins (Dob and Lec) which contain deletions corresponding to the entire hinge region of the heavy chains. Neither Dob nor Lec protein in aggregated form was able to activate the classical complement pathway, and this was shown to be due to an inability to bind the first component of complement (Cl). By rosette inhibition assays, Dob and Lec proteins were shown to have no measurable affinity for Fc receptors on human B cells or neutrophils. Dob and Lec proteins had a much reduced affinity for Fc receptors on the murine macrophage-like cell line P388D1 when compared to normal human IgGi. Furthermore, the hinge-deleted proteins were able to compete with murine IgG2b for P388D, receptors but not with murine IgG2a. In contrast, the binding of Dob and Lec proteins to protein A from Staphylococcus aureus was entirely normal. The functional consequences of the hinge deletion were parallel to those seen when normal IgGI was reduced and alkylated. It was concluded that the functional impotency of Dob and Lec proteins was related to the close association between the Fab and Fc regions in these molecules and the limited degree of segmental flexibility permitted in the absence of the hinge region. The data also suggest a major role for the C'y2 domain (C is the constant region) in mediating effector functions in normal IgGl.

Modulation of the Effector Functions of a Human IgG1 through Engineering of Its Hinge Region

“…The mechanisms by which the hinge could influence binding to C1q or Fc␥RIIIA include its inherent flexibility which allows the Fc to assume a favorable conformation or precludes the Fab arms from covering Fc interaction sites (5,13,33). Its role as a spacer preventing interference of the Fab arms with the Fc binding sites has also been suggested or discussed (1,3,13). However, as outlined in the introduction section, the exact nature of the relationship between the sequence and property of the hinge and the corresponding IgG effector functions is still to be determined.…”

The Solution Structures of Two Human IgG1 Antibodies Show Conformational Stability and Accommodate Their C1q and FcγR Ligands

“…Functionally, the reactivity of C1q with the four human IgG subclasses correlates with upper hinge length in the order of IgG3 Ͼ IgG1 Ͼ IgG2 Ͼ IgG4, with IgG4 not activating complement (3). A hingeless IgG1 antibody cannot bind or activate C1q (51). Mutagenesis studies of the hinge modulate C1q binding.…”

Crystallization and preliminary structure determination of an intact human immunoglobulin, b12: an antibody that broadly neutralizes primary isolates of HIV-1

“…While well over 200 structures of antibody fragments, mainly Fab and Fab H , have been determined, crystals of intact antibodies have only been reported eight times: Dob (Terry et al, 1968), Mcg (Edmundson et al, 1970), Kol (Palm & Colman, 1974), Zie (Ely et al, 1978), Mab231 (Larson et al, 1991;Harris et al, 1992), Mab4B7 (Stura et al, 1994), Mab24-404.1 and Mab61.1.3 (Harris et al, 1995 (Klein et al, 1981). The crystal structure determinations of Dob and Mcg revealed compact symmetrical planar T shapes (Silverton et al, 1977;Rajan et al, 1983;Guddat et al, 1993).…”