PEPTABODY - A NEW-TYPE OF HIGH AVIDITY BINDING-PROTEIN

A new type of high avidity binding molecule, termed ''peptabody'' was created by harnessing the effect of multivalent interaction, A short p eptide ligand was fused via a semi-rigid hinge region with the coiled- coil assembly domain of the cartilage oligomeric matrix protein, resul ting in a pentameric multivalent binding molecule. In the first peptab ody (Pab-S) described here, a peptide (S) specific for the mouse B-cel l lymphoma BCL(1) surface Ig idiotype, was selected from a phage displ ay library, A fusion gene was constructed encoding peptide S, followed by the 24 aa hinge region from camel Ige and a modified 55 aa cartila ge oligomeric matrix protein pentamerization domain. The Pab-S fusion protein was expressed in Escherichia coli in a soluble form at high le vels and purified in a single step by metal-affinity chromatography. P ab-S specifically bound the BCL(1) surface idiotype with an avidity of about 1 nM, which corresponds to a 2 x 10(5)-fold increase compared w ith the affinity of the synthetic peptide S itself. Biochemical charac terization showed that Pab-S is a stable homopentamer of about 85 kDa, with interchain disulfide bonds. Pab-S can be dissociated under denat uring and reducing conditions and reassociated as a pentamer with full -binding activity. This intrinsic feature provides an easy way to comb ine Pab molecules with two different peptide specificities, thus produ cing heteropentamers with bispecific acid/or chelating properties.