Cell Free Conversion of Designed Prion Proteins to Distinguish Ovine TSE Strains
Funded by DEFRA
Associated scientists: Dr. Louise Kirby, Mr. Will Brockie and Dr Andrew C. Gill
Distinguishing between TSE strains in vitro is not a facile process and the only reliable method involves passaging disease in a panel of laboratory mice. However, in the course of our research, we have developed an in vitro system that can model many aspects of TSE diseases in a test tube. The system is known as the cell free conversion assay (CFCA), and uses abnormal prion protein, from animals with TSE disease, to convert a synthetic prion protein that we have made in our laboratory. The process takes days, rather than months/years. We have also noticed that the conversion process proceeds more or less efficiently depending on the structure of the synthetic protein. Also, and crucially, conversion efficiency depends on the strain of TSE disease. This project aims to develop our cell free conversion assay system to result in a panel of synthetic proteins that will report on the identity of a TSE strain by assessing the extent to which each converts.
To develop our assay, we need to design the structures of the input panel of synthetic proteins to give maximum differentiation between TSE strains. To do this, we will chose natural protein variants that are already known or suspected to have some conversion differences between strains. We will also select certain areas of the prion protein for alteration to produce unnatural sequences not found in nature. For each TSE strain, we aim to produce one or two proteins that will convert with maximum efficiency, and use these proteins together in a panel that will result in a strain-specific pattern of conversion.
More details: please contact Andrew C. Gill