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Biotechnology and Applied Biochemistry (2003) 37, (173–182) (Printed in Great Britain)

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Concentration and removal of prion proteins from biological solutions

Brian Zeiler¹, Victor Adler, Valentin Kryukov and Abraham Grossman

Q-RNA, Inc., 3960 Broadway, New York, NY 10032-1543, U.S.A.

Key words: bovine spongiform encephalopathy (BSE), filtration, prion protein (PrP), RNA, RNA–protein interaction.

Abbreviations used: BSE, bovine spongiform encephalopathy; vCJD, variant Creutzfeldt-Jakob disease; PrP^C, normal cellular prion protein; PrP^Sc, abnormally folded isoform of PrP^C; mPrP, mouse PrP; hrPrP, human recombinant PrP^C protein; DTT, dithiothreitol; MBE, mouse brain extract; SELEX, systematic evolution of ligands by exponential enrichment.

¹To whom correspondence should be addressed (e-mail bzeiler@q-rna.com).

The pathogenesis of prion diseases is characterized by the accumulation of amyloid-like rods or scrapie-associated fibrils. The major protein component of scrapie-associated fibrils is an abnormally folded isoform of the normal cellular prion protein (PrP^C) that is resistant to digestion by proteinase K and is referred to as PrP^Sc. Purified human recombinant (hrPrP) was used to characterize the binding of a set of RNAs with affinity to PrP proteins. We report here that hrPrP has two RNA-binding activities at physiological pH. One activity is capable of binding all of the screened RNAs with high affinity, whereas the other activity can bind only to a subset of the RNAs with high affinity in the presence of non-specific competitor RNAs. A novel RNA belonging to the latter class, RQ11+12, bound to hrPrP with high affinity in the presence of vast molar excesses of competing RNAs. Beads impregnated with the RQ11+12 RNA were used to construct a filtration column. The column efficiently bound hrPrP and native PrP^C from serum and urine. Importantly, the filtration device was also capable of binding proteinase K-treated PrP^Sc from serum and urine. The level of sensitivity of detection of PrP by standard Western blotting was increased at least 1000-fold by first concentrating PrP from solution with the filtration column.

Received 27 September 2002/15 November 2002; accepted 29 November 2002

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