Three-dimensional structure of an anti-steroid Fab' and progesterone-Fab' complex.
The monoclonal anti-progesterone antibody DB3 binds progesterone with nanomolar affinity (Ka approximately 10(9) M-1), suggesting high specificity. However, DB3 also cross-reacts with similar affinity with a subgroup of structurally distinct, progesterone-like steroids. Crystals of the unliganded Fab' and various steroid-Fab' complexes are isomorphous and belong to the hexagonal space group, P6(4)22, with unit cell dimensions of a = b = 135 A, c = 124 A. Structures of free and progesterone-bound Fab' have been determined by X-ray crystallography at 2.7 A resolution using molecular replacement techniques. Progesterone is bound in a hydrophobic pocket formed mainly by the interaction of three complementarity determining regions L1, H2 and H3. The orientation of the ligand in the binding site was aided by both crystallographic and biochemical analyses of substituted steroids. The indole side-chain of TrpH100 of the DB3 has two different conformations, inter-converting "open" and "closed" forms of the antibody combining site. The TrpH100 indole thus appears to be acting as an antibody-derived surrogate ligand for its own hydrophobic binding pocket. These structures provide the first atomic view of how a steroid interacts with a protein and offer a structural explanation for the restriction of the anti-progesterone response to the VGAM3.8 family of VH genes.
Arevalo
J H
JH
Department of Molecular Biology, Scripps Research Institute, La Jolla, CA 92037.
Stura
E A
EA
Taussig
M J
MJ
Wilson
I A
IA
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000378:metabolism*; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D006825:Hybridomas / Q000276:immunology; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011374:Progesterone / Q000276:immunology / Q000378:metabolism*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013256:Steroids / Q000276:immunology*; D014961:X-Ray Diffraction
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D013256:Steroids; D011374:Progesterone