title: Characterization of conformation-dependent prion protein epitopes.
abstract: Whereas prion replication involves structural rearrangement of cellular prion protein (PrP(C)), the existence of conformational epitopes remains speculative and controversial, and PrP transformation is monitored by immunoblot detection of PrP(27-30), a protease-resistant counterpart of the pathogenic scrapie form (PrP(Sc)) of PrP. We now describe the involvement of specific amino acids in conformational determinants of novel monoclonal antibodies (mAbs) raised against randomly chimeric PrP. Epitope recognition of two mAbs depended on polymorphisms controlling disease susceptibility. Detection by one, referred to as PRC5, required alanine and asparagine at discontinuous mouse PrP residues 132 and 158, which acquire proximity when residues 126-218 form a structured globular domain. The discontinuous epitope of glycosylation-dependent mAb PRC7 also mapped within this domain at residues 154 and 185. In accordance with their conformational dependence, tertiary structure perturbations compromised recognition by PRC5, PRC7, as well as previously characterized mAbs whose epitopes also reside in the globular domain, whereas conformation-independent epitopes proximal or distal to this region were refractory to such destabilizing treatments. Our studies also address the paradox of how conformational epitopes remain functional following denaturing treatments and indicate that cellular PrP and PrP(27-30) both renature to a common structure that reconstitutes the globular domain.
authors.affiliation: Prion Research Center, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, Colorado 80523, USA.
authors.forename: Hae-Eun
authors.initials: HE
authors.lastname: Kang
authors.forename: Chu Chun
authors.initials: CC
authors.lastname: Weng
authors.forename: Eri
authors.initials: E
authors.lastname: Saijo
authors.forename: Vicki
authors.initials: V
authors.lastname: Saylor
authors.forename: Jifeng
authors.initials: J
authors.lastname: Bian
authors.forename: Sehun
authors.initials: S
authors.lastname: Kim
authors.forename: Laylaa
authors.initials: L
authors.lastname: Ramos
authors.forename: Rachel
authors.initials: R
authors.lastname: Angers
authors.forename: Katie
authors.initials: K
authors.lastname: Langenfeld
authors.forename: Vadim
authors.initials: V
authors.lastname: Khaychuk
authors.forename: Carla
authors.initials: C
authors.lastname: Calvi
authors.forename: Jason
authors.initials: J
authors.lastname: Bartz
authors.forename: Nora
authors.initials: N
authors.lastname: Hunter
authors.forename: Glenn C
authors.initials: GC
authors.lastname: Telling
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000096:biosynthesis / Q000302:isolation & purification; D002417:Cattle; D017124:Conserved Sequence; D003670:Deer; D019020:Directed Molecular Evolution; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D006801:Humans; D006825:Hybridomas; D051379:Mice; D008822:Mice, Transgenic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D018621:PrPC Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D018620:PrPSc Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012453:Saimiri; D017384:Sequence Deletion; D012756:Sheep
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D000939:Epitopes; D018621:PrPC Proteins; D018620:PrPSc Proteins; D011993:Recombinant Fusion Proteins