Quantifying the impact of membrane microtopology on effective two-dimensional affinity

Area of Science:

• Biophysics
• Cell Biology
• Immunology

Background:

• Membrane protein interactions are influenced by their surrounding environment, similar to soluble proteins.
• Adhesion receptors require close membrane apposition for function, making surface presentation critical.

Purpose of the Study:

• To demonstrate and quantify how distinct cellular environments affect the effective affinity and kinetic rates of adhesion receptors.
• To compare the binding characteristics of CD16b in red blood cells (RBCs), Chinese hamster ovary (CHO) cells, and K562 cells.

Main Methods:

• Engineered expression of CD16b (Fcgamma receptor) on RBCs, CHO cells, and K562 cells.
• Utilized a micropipette assay to measure effective affinity and kinetic rates of CD16b binding to IgG-coated surfaces.
• Employed electron microscopy to visualize cell-cell contacts.

Main Results:

• Receptors on RBCs exhibited a 50-fold higher effective affinity compared to those on CHO and K562 cells.
• Off-rates for CD16b were similar across all tested cell types.
• Electron microscopy showed broader contacts between RBC-RBC conjugates than CHO-RBC conjugates.

Conclusions:

• The cellular surface environment, particularly membrane presentation and roughness modulated by the cytoskeleton, significantly impacts adhesion molecule effectiveness.
• These findings have implications for understanding cell locomotion and static adhesion, even for receptors lacking cytoplasmic domains.