Screening a library of potential prion therapeutics against cellular prion proteins and insights into their mode of biological activities by surface plasmon resonance.
The conversion of cellular prion protein (PrP(C)) to the protease resistant isoform (PrP(Sc)) is considered essential for the progression of transmissible spongiform encephalopathies (TSEs). A potential therapeutic strategy for preventing the accumulation of PrP(Sc) is to stabilize PrP(C) through the direct binding of a small molecule to make conversion less energetically favourable. Using surface plasmon resonance (SPR)-based technology we have developed a procedure, based on direct binding, for the screening of small molecules against PrP(C) immobilized on a sensor chip. In this paper we report some problems associated with the immobilization of PrP(C) onto the sensor surface for conducting drug screening and how these problems were overcome. We demonstrated that the conformational change of PrP(C) on the chip surface leads to increased exposure of the C-terminal which was observed by the increase in quinacrine binding over time, and loss of heparin binding to the N-terminal. In addition, we also report the results of the successful screening of a library of 47 compounds of known activity in cell line or cell free conversion studies for direct binding to three forms of PrP(C) (huPrP(C), t-huPrP(C) and moPrP(C)). These results show the usefulness of this technique for the identification of PrP(C) binding ligands and to gain some insight as to their potential mode of action.
Touil
Faiza
F
The University of Sheffield, Department of Chemistry, Dainton Building, Brookhill, Sheffield S3 7HF, UK.
Pratt
Steven
S
Mutter
Roger
R
Chen
Beining
B
D015374:Biosensing Techniques; D002491:Central Nervous System Agents / Q000737:chemistry; D003224:Congo Red / Q000737:chemistry; D015195:Drug Design; D007700:Kinetics; D008024:Ligands; D010640:Phenothiazines / Q000737:chemistry; D018621:PrPC Proteins / Q000737:chemistry* / Q000378:metabolism; D018620:PrPSc Proteins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D020033:Protein Isoforms / Q000737:chemistry; D011796:Quinacrine / Q000737:chemistry; D020349:Surface Plasmon Resonance
D002491:Central Nervous System Agents; D008024:Ligands; D010640:Phenothiazines; D018621:PrPC Proteins; D018620:PrPSc Proteins; D020033:Protein Isoforms; D003224:Congo Red; D011796:Quinacrine