Structural and binding analysis of a two domain extracellular CD2 molecule.
The 50-kD CD2 (T11) surface glycoprotein on human T lymphocytes and thymocytes plays a critical role in T lineage cell activation and adhesion via its ligand LFA-3. To begin to define structure-function relationships in the extracellular segment of the transmembrane CD2 molecule, we have used a eukaryotic expression system and a CD2 cDNA to produce milligram amounts of recombinant soluble CD2 molecule that corresponds to the two extracellular segment exons. We show that this protein, termed T11ex2, behaves as a monomer in aqueous solution and includes a proteolytically resistant NH2-terminal fragment (domain I) encoded by the first extracellular segment exon. Circular dichroism analysis of T11ex2 demonstrates that its stabilized secondary structure is dependent on the intrachain disulfide bonds present in domain II. The T11ex2 monomer binds directly to the CD2 ligand LFA-3 with a dissociation constant of 0.4 microM. This relatively low affinity implies that cooperative binding resulting from an array of transmembrane CD2 molecules is important to facilitate physiologic T cell adhesion.
Sayre
P H
PH
Laboratory of Immunobiology, Dana-Farber Cancer Institute, Boston, Massachusetts.
Hussey
R E
RE
Chang
H C
HC
Ciardelli
T L
TL
Reinherz
E L
EL
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics; D000954:Antigens, Surface; D001402:B-Lymphocytes / Q000276:immunology; D018801:CD2 Antigens; D002460:Cell Line; D002942:Circular Dichroism; D003062:Codon; D004247:DNA / Q000235:genetics; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis; D005091:Exons; D006801:Humans; D007163:Immunosorbent Techniques; D008562:Membrane Glycoproteins; D010206:Papain / Q000378:metabolism; D010957:Plasmids; D011487:Protein Conformation; D011971:Receptors, Immunologic / Q000235:genetics; D011994:Recombinant Proteins; D012397:Rosette Formation; D013601:T-Lymphocytes / Q000276:immunology*; D014162:Transfection
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D018801:CD2 Antigens; D003062:Codon; D004220:Disulfides; D000939:Epitopes; D008562:Membrane Glycoproteins; D011971:Receptors, Immunologic; D011994:Recombinant Proteins; D004247:DNA; D010206:Papain