(331b) Motif-Grafted Domain Antibodies Selectively Recognize Amyloidogenic Conformers

Structure-function analysis of misfolded proteins associated with several conformational disorders (e.g., Parkinson’s and prion diseases) is greatly complicated by the fact that these proteins misfold into a spectrum of conformers with unique structures and biological activities. Conformation-specific antibodies that are immunoreactive with specific aggregated isoforms have been invaluable for isolating individual misfolded conformers for such structure-function studies, yet these antibodies are typically not sequence-specific and do not provide site-specific structural information. We have developed a motif-grafting strategy for generating small antibodies that are immunoreactive with misfolded proteins in a conformation- and sequence-specific manner. We find that grafting small hydrophobic peptide segments from multiple amyloidogenic proteins (including the Aβ peptide associated with Alzheimer’s disease) into the complementarity determining regions of a single domain antibody generates motif-grafted antibodies that selectively recognize soluble oligomers and amyloid fibrils of the corresponding amyloidogenic polypeptide. We also find that Aβ motif-grafted antibodies utilize homotypic interactions between grafted peptide segments and their cognate sequences within Aβ oligomers and fibrils to mediate specific recognition. Importantly, the motif-grafted antibodies for each polypeptide are highly sequence specific, as they do not cross react with fibrils of other amyloidogenic proteins and scrambling the grafted sequences eliminates their immunoreactivity. We expect that the simplicity of synthesizing libraries of motif-grafted antibodies will facilitate their use in investigating conformational differences between aggregated isoforms for diverse misfolded proteins.