Immunoglobulin Binding Specificities of the Homology Regions (Domains) of Protein A

Abstract

Protein A binds immunoglobulins and it has two target structures, one in Fc gamma (CH) and the other in selected VH regions. The protein has five homology regions (domains), A, B, C, D, and E. Fc-binding and VH-binding have been reported to be non-competitive, suggesting that different domains are responsible for the binding of the two ligands. On the other hand, all five domains have been reported to bind Fc. I studied binding of different immunoglobulins by protein A or its domain B (rBB). The results show that separate domains bind VH and Fc. If all five domains are capable of binding Fc, the ones that bind VH have low affinity for Fc. Furthermore, the number of Fc-binding domains varies depending on the type of the IgG being bound. Human IgG1 or IgG2 or rabbit IgG (Fc) seem to be bound by several domains (possibly four), and domain B is one of them. Mouse IgG1 or IgG2b are bound by fewer domains not including B. Murine IgG2a is also bound by fewer domains but B is one of them.

All individual domains of staphylococcal protein A show Fab binding

“…Previously, domains E and/or D have been suggested to be responsible for the main Fab binding of protein A [8, 9, 42]. Our data does not support this hypothesis as domains A, B, and C in our experiments all bind Fab with similar strength as domains E and D. In conclusion, this work has demonstrated that all five native SPA domains E, D, A, B and C show affinity for both Fab and Fc.…”

“…However, the binding analysis of separate monovalent protein A fragments to F(abP) P and to the scFv gave the unexpected result that domain B clearly binds Fab whereas the very similar domain Z shows only little or no binding activity. Although the observed weak binding for domain Z to Fab is in accordance with earlier results [40], a stronger Fab binding of domain B is observed in this study as compared to precious ¢ndings [8,40]. However, data such as shown in Fig.…”

“…To date, there have been limited efforts to analyse the relative binding strengths of the five individual homologous SPA domains towards Fab and Fc, respectively. It has been indicated that not all domains are involved in the Fab interaction [8]. Recently, it was demonstrated that domain D binds to Fab, but the other individual domains were not included in the study [9].…”

In vitro Selection and Interaction Studies of a DNA Aptamer Targeting Protein A

“…A relatively weak binding signal was observed for the human Fab fragment. Less common is the ability of Protein A to bind the Fab fragment of immunoglobulins, especially the Fab heavy chain V H 3 family [ 7 , 8 , 43 , 44 ]. It was found that both Fc and Fab fragments bind to Protein A in a noncompetitive way and that they use the same Ig-binding domains, but different epitopes inside of these domains [ 44 ].…”

Staphylococcal Protein A Binding to VH3Encoded Immunoglobulins

“…This suggests that domains A, D and/or E contain at least one site necessary for Fab binding. In one study rBB exhibited binding to an isolated recombinant VH3 [17], whereas in another study, the same recombinant fragment B failed to bind five monoclonal antibodies bound by SPA via the VH region [28]. The basis for these different findings remains unresolved.…”