CHARACTERIZATION OF THE LOW-AFFINITY INTERACTION BETWEEN RAT-CELL ADHESION MOLECULES CD2 AND CD48 BY ANALYTICAL ULTRACENTRIFUGATION

CD2 is a cell adhesion molecule found on the plasma membrane of T-lymp hocytes. Its counter-receptor in rat is the structurally related CD48. This interaction is believed to contribute to the adhesion of T-cells to other cells such as cytotoxic targets and antigen presenting cells . Cell-cell adhesion involves the formation of multiple cell adhesion molecule complexes at the cell surface and if cell-cell de-adhesion is to occur, these complexes need to be disrupted. The affinities of cel l adhesion molecule interactions are suggested to be relatively weak t o allow this de-adhesion of cell-cell interactions. The CD2/CD48 inter action has been studied using recombinant extracellular proteins and t he affinity of the interaction of soluble recombinant rat CD2-CD48 has been determined (at 37 degrees C) using surface plasmon resonance (an d shown to be weak), with the dissociation constant K-d = 60-90 mu M. The values determined by surface plasmon resonance results could be af fected by the immobilisation of the ligand on the chip and any self-as sociation on the chip. We used three different analytical ultracentrif uge procedures which each allowed the interaction to be studied in fre e solution without the need for an immobilisation medium. Both sedimen tation equilibrium (using direct analysis of the concentration distrib ution and also modelling of molecular weight versus concentration data ) and sedimentation velocity at 5 degrees C yielded dissociation const ants in the range of 20-110 mu M, supporting the surface plasmon reson ance findings showing that binding between these cell adhesion molecul es is relatively weak. These studies also ruled out the presence of an y significant self-association of the reactants which could lead to sy stematic error in the surface plasmon resonance results.