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Journal of Virology, December 2000, p. 11928-11934, Vol. 74, No. 24
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Affinity-Tagged Miniprion Derivatives Spontaneously Adopt Protease-Resistant Conformations

Surachai Supattapone,1,2 Hoang-Oanh B. Nguyen,1,2 Tamaki Muramoto,1,dagger Fred E. Cohen,1,2,3,4 Stephen J. DeArmond,1,5 Stanley B. Prusiner,1,2,4,* and Michael Scott1,2

Institute for Neurodegenerative Diseases1 and Departments of Neurology,2 Cellular and Molecular Pharmacology,6 Medicine,3 Pathology,5 and Biochemistry and Biophysics,4 University of California at San Francisco, San Francisco, California 94143

Received 14 July 2000/Accepted 22 September 2000

An abridged PrP molecule of 106 amino acids designated PrP106 can form infectious miniprions in transgenic (Tg) mice (29). Addition of six-histidine (His6) affinity tags to selective sites within PrP106 resulted unexpectedly in new PrP proteins that spontaneously adopted protease-resistant conformations when expressed in neuroblastoma cells and Tg mice. Acquisition of protease resistance depended on the length, charge, and placement of the affinity tag. Introduction of the disease-linked mutation E200K into the sequence of PrP106(140/6His) increased the recovery of protease-resistant PrP fivefold, whereas introduction of the mutations C213A and Delta 214-220 did not affect the recovery of protease-resistant PrP. Treatment of cultured cells expressing affinity-tagged PrP106 mutants with polypropyleneimine dendrimer rendered these proteins sensitive to protease digestion in a manner similar to wild-type PrPSc. We conclude that certain affinity-tagged PrP106 proteins spontaneously fold into conformations partially resembling, yet distinct from, wild-type PrPSc. These proteins might be useful tools in the identification of new disease-causing mutations as well as for screening compounds for therapeutic efficacy.

* Corresponding author. Mailing address: Institute for Neurodegenerative Diseases, Box 0518, University of California, San Francisco, CA 94143-0518. Phone: (415) 476-4482. Fax: (415) 476-8386. E-mail: ind{at}itsa.ucsf.edu.

dagger Present address: Tohoku University School of Medicine, 2-1 Seiryou-Machi, Aoba-ku, Sendai, 980 Japan.

Journal of Virology, December 2000, p. 11928-11934, Vol. 74, No. 24
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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