Molecular analysis of IgM rheumatoid factor binding to chimeric IgG

Area of Science:

• Immunology
• Structural Biology
• Autoimmunity

Background:

• Rheumatoid factors (RF) are autoantibodies that target immunoglobulin G (IgG), playing a role in autoimmune diseases like rheumatoid arthritis.
• The binding specificities of RF to different IgG subclasses are not fully understood, impacting disease pathogenesis and diagnostic strategies.

Purpose of the Study:

• To identify the specific regions on IgG responsible for binding by IgM RF.
• To elucidate the role of IgG constant region domains and glycosylation in RF recognition.

Main Methods:

• Utilized chimeric IgG antibodies with shuffled murine and human constant regions.
• Employed a modified RF ELISA to assess binding of polyclonal and monoclonal RF to various IgG subclasses.
• Generated aglycosylated IgG antibodies by mutating the Asn-297 residue to investigate the role of N-linked carbohydrates.

Main Results:

• Polyclonal RF from rheumatoid arthritis patients bound IgG1, IgG2, and IgG4 strongly, with weaker binding to IgG3.
• Monoclonal RF from Waldenstrom's macroglobulinemia patients predominantly bound IgG1, IgG2, and IgG4, with limited reactivity to IgG3.
• The CH3 domain of IgG was identified as the primary determinant for differential RF binding between IgG3 and IgG4.
• Aglycosylated IgG3 showed enhanced binding by RF compared to glycosylated IgG3, suggesting a role for glycosylation in IgG3 recognition.

Conclusions:

• The CH3 domain variation significantly influences monoclonal RF binding specificity across IgG subclasses.
• While glycosylation generally does not affect RF binding to IgG1, IgG2, and IgG4, it plays a role in IgG3 recognition.
• These findings provide critical insights into the structural basis of RF-IgG interactions and their implications in autoimmune diseases.