NR ATQW
AU Schaller,M.; Solforosi,L.; Moroncini,G.; Abalos,G.C.; Cruite,J.T.; Williamson,R.A.
TI Molecular characterization of motif-grafted PrPsc-specific antibodies
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Structure of PrP and molecular determinants of infectivity STRCT-12
PT Konferenz-Poster
AB
Antibody scaffolds containing grafted polypeptides comprised either of PrP amino acid residues 89-112 or of PrP residues 136-158, bind specifically and with high affinity, to PrPsc and PrP27-30 from prion-infected humans and rodents, but not to PrPc. Characterizing the nature of the specific binding interaction between the motif-grafted antibody reagents and disease-associated PrP conformers may yield insights into the formation of the PrPc-PrPsc interface, a key step in the assembly of the prion replicative complex.
Binding experiments indicate that the PrPsc epitope recognized by IgG 136-158 is proteinaceous, whereas recognition of PrPsc and PrP27-30 in brain homogenates by IgG 89-112 was diminished by pre-treatment of the homogenate with DNase, but not RNase. Moreover, binding of IgG 89-112 to the abnormal PrP conformers was readily inhibited in the presence of excess DNA. These observations suggest that binding to PrPsc and PrP27-30 by IgG 89-112 is at least partially dependent upon the presence of DNA, implying the existence of a complex containing PrPsc and DNA in homogenates of prion-infected brains. To identify key residues within the 89-112 PrP sequence graft contributing to the PrPsc binding interaction, we have generated an additional series of antibodies containing truncated and mutated PrP grafts. Binding studies using these reagents have identified a core PrPsc binding motif composed of PrP amino acids 95-105, within which lysine residues at positions 101 and 104 appear to perform a critical role. In addition, to determine the structure of the grafted PrP sequence within the antibody scaffold, crystal structures of Fab fragments of IgGs 89-112 and 136-158 are presently being resolved.
An extensive panel of novel motif-grafted antibodies containing other regions of PrP peptide sequence have been generated to identify possible additional sites of interaction between PrPc and PrPsc.
AD Monica Schaller, Laura Solforosi (lsolf@scripps.edu), Gianluca Moroncini, Gil C. Abalos III, Justin T. Cruite, R. Anthony Williamson (anthony@scripps.edu), Department of Immunology, The Scripps Research Institute, La Jolla, California 92037, USA; Gianluca Moroncini, University of Ancona, Italy
SP englisch
PO Deutschland