Serial protein pairs in cell/surface adhesion

Abstract

The theoretical and experimental work described offers the first systematic investigation of cell/surface adhesion mediated through protein/protein bonds in series. Cell adhesion and migration are facilitated through protein/protein bonds in series established when membrane-bound proteins simultaneously interact with extracellular proteins and with intracellular cytoskeletal components. The biotechnological application of affinity-based cell separation exploits serial protein/protein bonds to isolate a target cell type from a heterogeneous pool of cells.

A mathematical model describing the attachment and detachment of protein-coated cells through soluble linker proteins to a protein-coated substrate is developed following a deterministic, mass-action approach. The key prediction is that the force to rupture cell/substrate contacts comprised of protein/protein bonds in series is always less than the force to rupture contacts of either of the bonds separately. Also, the detachment force is a maximum over a narrow range of linker protein concentration. The distribution of linkage fracture location during cell detachment has only a mild dependence on the ratio of the bond affinities.

A model system is selected which uses protein-coated beads for cells, protein-coated glass plates for biological substrates, and three proteins, protein A, human IgG, and anti-human IgG. The Radial-Flow Detachment Assay is employed to measure the specific adhesion strength of protein/protein interactions for bead/plate contacts connected by single, protein A/human IgG interactions; single, human IgG/anti-human IgG interactions; serial, protein A/human IgG/anti-human IgG interactions; and background, protein A/anti-human IgG interactions. Experimental results confirm that the force to rupture cell/substrate contacts comprised of protein/protein bonds in series is less than the force to rupture contacts of either of the bonds separately. The adhesion strength of the contacts mediated through serial linkages is different from the background interaction and is sensitive to the soluble linker concentration.

The model predictions and experimental results are synthesized to provide suggestions to improve the selective retention of target cells during an affinity-based cell separation.