The increased potency of cross-linked lymphocyte function-associated antigen-3 (LFA-3) multimers is a direct consequence of changes in valency.
We have used chemically cross-linked dimers, trimers, and tetramers of lymphocyte function-associated antigen-3 (LFA-3) to study the role of multivalency in the interaction of the protein with its receptor, CD2. The cross-linked adducts showed enhanced activity in systems where LFA-3 has been shown to (i) block LFA-3/CD2 interactions in a rosetting assay and (ii) provide through the CD2 on peripheral blood lymphocytes a trigger for T-cell proliferation. The level of increase was directly related to the valency state of the multimers. In the rosetting assay, the dimers, trimers, and tetramers, by weight, exhibited 15-, 150-, and 430-fold increases in activity over monomeric LFA-3. In the proliferation assay, the tetramer produced a 6-fold increase in thymidine incorporation at 0.06 micrograms/ml, the trimer was 100 times less active than the tetramer, and the dimer and monomer were inactive. The LFA-3 multimers were generated using a three-step cross-linking chemistry that was targeted at the carbohydrates on LFA-3. With this procedure over 60% of the starting protein was converted into multimers with no effect on function. The cross-linking approach should be applicable to other surface antigens, providing a simple method for analyzing multivalent interactions.
Pepinsky
R B
RB
Biogen, Inc., Cambridge, Massachusetts 02142.
Chen
L L
LL
Meier
W
W
Wallner
B P
BP
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D000945:Antigens, Differentiation, T-Lymphocyte / Q000378:metabolism; D000954:Antigens, Surface / Q000737:chemistry*; D018801:CD2 Antigens; D018968:CD58 Antigens; D006224:Cricetinae; D003412:Cricetulus; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000737:chemistry; D006801:Humans; D008213:Lymphocyte Activation; D008562:Membrane Glycoproteins / Q000737:chemistry*; D011971:Receptors, Immunologic / Q000378:metabolism; D012397:Rosette Formation; D013601:T-Lymphocytes / Q000737:chemistry / Q000276:immunology
D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D018801:CD2 Antigens; D018968:CD58 Antigens; D003432:Cross-Linking Reagents; D008562:Membrane Glycoproteins; D011971:Receptors, Immunologic