Misfolded protein sensor method in body fluids

Abstract

A catalytic conformational sensor method for detecting abnormal proteins and proteinaceous particles. The method is based on the interaction of a peptide fragment or probe with an abnormal proteinaceous particle. The interaction catalyzes transformation of the probe to a predominately beta sheet conformation and allows the probe to bind to the abnormal proteinaceous particle. This in turn, catalyzes propagation of a signal associated with the test sample-bound probe. As a result signals can be propagated even from samples containing very low concentrations of abnormal proteinaceous particles as is the case in many body-fluid derived samples.

1 .- 20 . (canceled)  
     
     
         21 . An isolated peptide reagent that interacts preferentially with pathogenic forms of a conformational disease protein as compared to nonpathogenic forms of the conformational disease protein.  
     
     
         22 . The peptide reagent of  claim 21 , wherein the peptide reagent includes the amino acid sequence (G)n, where n=1, 2, 3 or 4, at the N-terminal end, at the C-terminal end, or at both the N-terminal and C-terminal end.  
     
     
         23 . The peptide reagent of  claim 21 , wherein the peptide reagent is genetically encoded.  
     
     
         24 . A polynucleotide encoding a peptide reagent according to  claim 23 .  
     
     
         25 . A composition comprising the polynucleotide of  claim 24 .  
     
     
         26 . The peptide reagent of  claim 21 , wherein the conformational disease is a prion-related disease, the pathogenic protein is PrP Sc , and the nonpathogenic form is PrP C .  
     
     
         27 . The peptide reagent of  claim 26 , wherein the peptide reagent is derived from a fragment of a prion protein.  
     
     
         28 . A composition comprising a peptide reagent according to  claim 26 .  
     
     
         29 . A complex comprising the peptide reagent of  claim 26  and a pathogenic prion protein.  
     
     
         30 . A peptide having a predominantly alpha-helix secondary structure, random coil secondary structure, or a combination thereof, that interacts with misfolded proteinaceous particles.  
     
     
         31 . The peptide of  claim 30 , wherein the misfolded proteinaceous particles are PrP SC  particles.  
     
     
         32 . The peptide of  claim 30 , wherein the peptide undergoes a conformational shift that results in a decrease in alpha-helix and/or random coil secondary structure and an increase in beta-sheet secondary structure upon contact with misfolded proteinaceous particles or upon contact with another such peptide that has undergone such a conformational shift.  
     
     
         33 . The peptide of  claim 30 , wherein the peptide has a helix-loop-helix structure.  
     
     
         34 . The peptide of  claim 30 , wherein the peptide comprises a sequence found in a wild-type transmissibile spongiform encephalopathy (TSE) peptide sequence, a species-specific TSE peptide sequence, or a mutated TSE sequence mutatet to be destabilized and/or noninfectious.  
     
     
         35 . The peptide of  claim 30 , wherein the peptide is labeled with a detectable label.  
     
     
         36 . The peptide of  claim 30 , wherein the peptide is non-infectious.  
     
     
         37 . A composition comprising a peptide of  claim 30  bound to a misfolded proteinaceous particle.  
     
     
         38 . A composition comprising a peptide of  claim 30 .  
     
     
         39 . The composition of  claim 38 , wherein the misfolded proteinaceous particle is a PrP SC  particle.  
     
     
         40 . A method for detecting the presence of a pathogenic prion in a sample comprising: 
 (a) contacting a sample suspected of comprising a pathogenic prion with a first peptide reagent according to  claim 26  under conditions that allow binding of the first peptide reagent to the pathogenic prion protein, if present; and    (b) detecting the presence the pathogenic prion, if any, in the sample by its binding to the first peptide reagent.    
     
     
         41 . The method of  claim 40  wherein the first peptide reagent is detectably labeled.  
     
     
         42 . A method for detecting the presence of a pathogenic prion in a sample comprising: 
 (a) contacting a sample suspected of comprising a pathogenic prion with a first peptide reagent according to  claim 26  under conditions that allow interaction of the first peptide reagent with the pathogenic prion protein, if present; and    (b) detecting the presence the pathogenic prion, if any, in the sample by its interaction with the first peptide reagent.    
     
     
         43 . A method for detecting the presence of a pathogenic prion in a sample comprising: 
 (a) contacting a sample suspected of comprising a pathogenic prion with a first peptide reagent according to  claim 30  under conditions that allow interaction of the first peptide reagent to the pathogenic prion protein, if present; and    (b) detecting the presence the pathogenic prion, if any, in the sample by its interaction with the first peptide reagent.