A molecular model of RGD ligands. Antibody D gene segments that direct specificity for the integrin alpha IIb beta 3.
Following an ill-defined activation event, the Arg-Gly-Asp (RGD) recognition site of the platelet integrin, glycoprotein IIb-IIIa (alpha IIb beta 3), can bind to fluid-phase, RGD-containing protein ligands, such as fibrinogen, or to the murine monoclonal IgM, PAC-1, which contains the sequence Arg-Tyr-Asp (RYD) within the third complementarity-determining region of its heavy chain (H3). PAC-1 has thus become a widely exploited marker of platelet alpha IIb beta 3 activation. In this report, we compare PAC-1 with two murine IgG, OP-G2 (IgG1 kappa) and LJ-CP3 (IgG1 kappa), that also contain the sequence RYD in H3 but bind to alpha IIb beta 3 without prior activation. Each antibody can inhibit the binding of the other two to intact platelets or to purified IIb-IIIa, the binding of each antibody is completely inhibited by peptides containing RGD, and H3 of each antibody uses the germline D-gene DSP 2.10 (CTATAGGTACGAC) which includes the sequence RYD. Two other murine IgG, HP20 and PCG1-1, cloned and sequenced by other laboratories, also utilize the DSP 2.10 sequence, but neither antibody binds to alpha IIb beta 3. From a comparison of the H3 sequences of these antibodies, we have developed a molecular model of the H3 loop region which can explain these differences in specificity. This model predicts that both the ability to bind to alpha IIb beta 3 and the activation dependence of that binding are a function of the orientation and, therefore, accessibility of the RYD sequence. This model and refinements thereof can be exploited to study the molecular basis for specificity and affinity of RGD-containing ligands for integrins.
Tomiyama
Y
Y
Blood Research Institute of the Blood Center of Southeastern Wisconsin, Milwaukee 53233.
Brojer
E
E
Ruggeri
Z M
ZM
Shattil
S J
SJ
Smiltneck
J
J
Gorski
J
J
Kumar
A
A
Kieber-Emmons
T
T
Kunicki
T J
TJ
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000918:Antibody Specificity; D001483:Base Sequence; D001792:Blood Platelets / Q000378:metabolism*; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007145:Immunoglobulin kappa-Chains / Q000235:genetics / Q000378:metabolism; D016023:Integrins / Q000276:immunology / Q000378:metabolism*; D008024:Ligands; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D009842:Oligopeptides / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid; D014961:X-Ray Diffraction
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains; D016023:Integrins; D008024:Ligands; D009838:Oligodeoxyribonucleotides; D009842:Oligopeptides; D010980:Platelet Membrane Glycoproteins; C047981:arginyl-glycyl-aspartic acid