Ionic strength and transition metals control PrPSc protease resistance and conversion-inducing activity.
The essential component of infectious prions is a misfolded protein termed PrPSc, which is produced by conformational change of a normal host protein, PrPC. It is currently unknown whether PrPSc molecules exist in a unique conformation or whether they are able to undergo additional conformational changes. Under commonly used experimental conditions, PrPSc molecules are characteristically protease-resistant and capable of inducing the conversion of PrPC molecules into new PrPSc molecules. We describe the effects of ionic strength, copper, and zinc on the conformation-dependent protease resistance and conversion-inducing activity of PrPSc molecules in scrapie-infected hamster brains. In the absence of divalent cations, PrPSc molecules were > 20-fold more sensitive to proteinase K digestion in low ionic strength buffers than in high ionic strength buffers. Addition of micromolar concentrations of copper or zinc ions restored the protease resistance of PrPSc molecules under conditions of low ionic strength. These transition metals also controlled the conformation of purified truncated PrP-(27-30) molecules at low ionic strength, confirming that the N-terminal octapeptide repeat region of PrPSc is not required for binding to copper or zinc ions. The protease-sensitive and protease-resistant conformations of PrPSc were reversibly interchangeable, and only the protease-resistant conformation of PrPSc induced by high ionic strength was able to induce the formation of new protease-resistant PrP (PrPres) molecules in vitro. These findings show that PrPSc molecules are structurally interconvertible and that only a subset of PrPSc conformations are able to induce the conversion of other PrP molecules.
Nishina
Koren
K
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
Jenks
Samantha
S
Supattapone
Surachai
S
D000818:Animals; D001921:Brain / Q000378:metabolism; D002412:Cations; D002462:Cell Membrane / Q000378:metabolism; D003300:Copper / Q000737:chemistry; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D019286:Endopeptidase K / Q000494:pharmacology; D010450:Endopeptidases / Q000737:chemistry; D015151:Immunoblotting; D007477:Ions; D008647:Mesocricetus; D008670:Metals; D051379:Mice; D018620:PrPSc Proteins / Q000737:chemistry / Q000494:pharmacology*; D011487:Protein Conformation; D012608:Scrapie / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors; D015032:Zinc / Q000737:chemistry
D002412:Cations; D007477:Ions; D008670:Metals; D018620:PrPSc Proteins; D003300:Copper; D010450:Endopeptidases; D019286:Endopeptidase K; D015032:Zinc