EXPLORING THE MIMICRY OF POLYSACCHARIDE ANTIGENS BY ANTIIDIOTYPIC ANTIBODIES - THE CRYSTALLIZATION, MOLECULAR REPLACEMENT, AND REFINEMENT TO 2-CENTER-DOT-8 ANGSTROM RESOLUTION OF AN IDIOTOPE-ANTI-IDIOTOPE FAB COMPLEX AND OF THE UNLIGANDED ANTI-IDIOTOPE FAB

The monoclonal antibody YsT9.1 is specific for the lipopolysaccharide A antigen of Brucella abortus. A complex formed between the Fab of YsT 9.1 (Ab1) and the Fab of its anti-idiotopic monoclonal antibody T91AJ5 (Ab2) has been crystallized, and data hare been collected to 2.8 Angs trom resolution. The space group is monoclinic P2(1), with one molecul e per asymmetric unit. The structure was solved using a limited Patter son-correlation search over the asymmetric-unit of rotation space, and has been refined to an R-factor of 0.174. The complex is a head-to-he ad dimer, with the contact between the two Fabs almost completely rest ricted to their hypervariable loops. The interactions between the two Fabs in the complex are dominated by tyrosine residues, not only in th e formation of hydrogen bonds, but in their participation in an aromat ic ring network that spans the two Fv domains. The anti-idiotope was f ound to be unable to carry an internal image of the antigen and induce polysaccharide-specific ''anti-anti-idiotopes'' (Ab3) because the pol ysaccharide binding cleft on the Ab1 is too narrow and deep to allow c omprehensive contact with the binding site of Ab2. The antibody T91AJ5 therefore is a class gamma anti-idiotope. The contact surfaces of the two Fabs are highly complementary; however, they are distinctly diffe rent in character. Each Fab has two separate binding surfaces of appro ximately equal size? but while the two binding surfaces of Ab1 are par titioned between the light and heavy chains, the two binding surfaces of Ab2 are shared between the chains, with the heavy chain responsible for about 60% of the total binding area. The structure of the unligan ded Fab of Ab2 has also been solved by molecular replacement, and refi ned to an R-factor of 0.152 at 2.8 Angstrom resolution. This Fab cryst allizes in the orthorhombic space group P2(1)2(1)2(1), with one molecu le per asymmetric unit. The second hypervariable loop of the heavy cha in of the Ab2 Fab is observed to undergo a significant and necessary c onformational rearrangement in going from the unliganded to complexed form, and thus complex formation is an example of ''induced fit'' of a ntigen to antibody. The unliganded Ab1 Fab packs in the standard head- to-tail fashion observed for other Fabs. This mode of packing is precl uded in the complex, by its head-to-head nature, and it is found to pa ck in tilted layers with most intermolecular contacts made between adj acent Ab2 Fab molecules. The unliganded Ab2 Fab packs in a manner that cannot be described as either head-to-tail or head-to-head.