Prion Protein Isoforms, a Convergence of Biological and Structural Investigations (1995)

Prion diseases are fatal neurodegenerative disorders afflicting humans and other mammals. Manifest as sporadic, inherited, and infectious illnesses, they seem to be the result of a conformational transition in the normal, cellular prion protein (PrPC) 1The abbreviations used are:PrPprion proteinBSEbovine spongiform encephalopathyCJDCreutzfeldt-Jakob diseaseGSSGerstmann-Sträussler-Schienker syndromeFFIfatal familial insomniaGPIglycosylphosphatidylinositolFTIRFourier transform infraredTgtransgenic. whereby the aberrant, diseased protein isoform (PrPSc) is formed(1Prusiner S.B. Science. 1991; 252: 1515-1522Google Scholar). The transmissible particle or prion is composed largely, if not exclusively, of PrPSc and is resistant to procedures that modify nucleic acids(2Alper T. Cramp W.A. Haig D.A. Clarke M.C. Nature. 1967; 214: 764-766Google Scholar, 3Prusiner S.B. Science. 1982; 216: 136-144Google Scholar). No candidate scrapie-specific polynucleotide has been identified(4Kellings K. Meyer N. Mirenda C. Prusiner S.B. Riesner D. J. Gen. Virol. 1992; 73: 1025-1029Google Scholar). The name prion was coined for the proteinaceous infectious agent (3Prusiner S.B. Science. 1982; 216: 136-144Google Scholar), and the designation PrPSc was derived from scrapie, an infectious disease of sheep known for at least 200 years that has been transmitted to hamsters and mice(5Chandler R.L. Lancet. 1961; 1: 1378-1379Google Scholar, 6Marsh R.F. Kimberlin R.H. J. Infect. Dis. 1975; 131: 104-110Google Scholar). PrPC and PrPSc have been purified from the brains of healthy and scrapie-infected Syrian hamsters, respectively. Among a plethora of early proposals as to the possible causes of scrapie were suggestions of a protein devoid of nucleic acid(2Alper T. Cramp W.A. Haig D.A. Clarke M.C. Nature. 1967; 214: 764-766Google Scholar, 7Pattison I.H. Gajdusek D.C. Gibbs Jr., C.J. Alpers M.P. Slow, Latent and Temperate Virus Infections, NINDB Monograph 2. U. S. Government Printing Office, Washington, D. C.1965: 249-257Google Scholar, 8Pattison I.H. Jones K.M. Vet. Rec. 1967; 80: 1-8Google Scholar, 9Griffith J.S. Nature. 1967; 215: 1043-1044Google Scholar). Over the last two decades, substantial evidence has been gathered suggesting that PrPSc is likely to be the only component of the infectious prion particle. prion protein bovine spongiform encephalopathy Creutzfeldt-Jakob disease Gerstmann-Sträussler-Schienker syndrome fatal familial insomnia glycosylphosphatidylinositol Fourier transform infrared transgenic. More than 150,000 cattle have died of a new prion disease called bovine spongiform encephalopathy (BSE) or “mad cow disease”(10Wilesmith J.W. Prusiner S.B. Collinge J. Powell J. Anderton B. Prion Diseases of Humans and Animals. Ellis Horwood Ltd., London1992: 243-255Google Scholar), believed to have originated from cattle feed infected with sheep scrapie prions. A 1988 ban on the feed produced from the offal of domestic animals has begun to control the epidemic; in 1992, ~37,000 cases of BSE were recorded while only ~22,000 were diagnosed in 1994. Prion diseases of humans include Creutzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Schienker syndrome (GSS), and fatal familial insomnia (FFI)(11Gajdusek D.C. Science. 1977; 197: 943-960Google Scholar, 12Masters C.L. Gajdusek D.C. Gibbs Jr., C.J. Brain. 1981; 104: 559-588Google Scholar, 13Medori R. Tritschler H.-J. LeBlanc A. Villare F. Manetto V. Chen H.Y. Xue R. Leal S. Montagna P. Cortelli P. Tinuper P. Avoni P. Mochi M. Baruzzi A. Hauw J.J. Ott J. Lugaresi E. Autilio-Gambetti L. Gambetti P. N. Engl. J. Med. 1992; 326: 444-449Google Scholar). Generally patients with GSS present with ataxia and cerebellar dysfunction while those with CJD exhibit memory loss and a progressive decline of intellect. GSS and FFI are familial diseases whereas CJD can be sporadic, familial, or infectious. The infectious human prion diseases are rare, but kuru spread through ritualistic cannibalism was once the most common fatal illness among the Fore people of Papua New Guinea(11Gajdusek D.C. Science. 1977; 197: 943-960Google Scholar). Iatrogenic CJD is another infectious prion disease resulting from inadequately sterilized surgical instruments, dura mater grafts, and human growth hormone isolated from the pituitaries of cadavers. Prion diseases vary in their course and rapidity of development, but they all cause neurodegeneration and ultimately death. Autopsy generally reveals spongiform degeneration of the brain, sometimes with amyloid plaques that stain with Congo red and react with α-PrP antibodies. The protease-resistant fragment of PrPSc called PrP 27-30 was discovered in fractions partially purified from Syrian hamster (SHa) brain by monitoring the enrichment of scrapie infectivity with respect to protein(14Prusiner S.B. Bolton D.C. Groth D.F. Bowman K.A. Cochran S.P. McKinley M.P. Biochemistry. 1982; 21: 6942-6950Google Scholar, 15Bolton D.C. McKinley M.P. Prusiner S.B. Science. 1982; 218: 1309-1311Google Scholar, 16Prusiner S.B. Cochran S.P. Groth D.F. Downey D.E. Bowman K.A. Martinez H.M. Ann. Neurol. 1982; 11: 353-358Google Scholar). Differential solubilization with detergents, limited proteolysis to remove extraneous proteins and centrifugation were used to purify scrapie prions. The purification was extended using discontinuous sucrose gradient centrifugation. PrP 27-30 aggregates to form amyloid rods that stain with Congo red dye to display green-gold birefringence under polarized light(17Prusiner S.B. McKinley M.P. Bowman K.A. Bolton D.C. Bendheim P.E. Groth D.F. Glenner G.G. Cell. 1983; 35: 349-358Google Scholar). N-terminal sequencing of PrP 27-30 revealed a ragged N terminus(18Prusiner S.B. Groth D.F. Bolton D.C. Kent S.B. Hood L.E. Cell. 1984; 38: 127-134Google Scholar). The N-terminal amino acid sequence allowed the selection of a clone from a hamster brain cDNA library and the identification of the PrP gene (19Chesebro B. Race R. Wehrly K. Nishio J. Bloom M. Lechner D. Bergstrom S. Robbins K. Mayer L. Keith J.M. Garon C. Haase A. Nature. 1985; 315: 331-333Google Scholar, 20Oesch B. Westaway D. Wälchli M. McKinley M.P. Kent S.B.H. Aebersold R. Barry R.A. Tempst P. Teplow D.B. Hood L.E. Prusiner S.B. Weissmann C. Cell. 1985; 40: 735-746Google Scholar). Subsequently, both PrPC and PrPSc were found to be encoded within a single exon of the chromosomal gene as proteins of 254 amino acids(21Basler K. Oesch B. Scott M. Westaway D. Wälchli M. Groth D.F. McKinley M.P. Prusiner S.B. Weissmann C. Cell. 1986; 46: 417-428Google Scholar), although they display quite different physical properties(22Meyer R.K. McKinley M.P. Bowman K.A. Braunfeld M.B. Barry R.A. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1986; 83: 2310-2314Google Scholar). N-terminal sequencing of full-length PrPC(23Turk E. Teplow D.B. Hood L.E. Prusiner S.B. Eur. J. Biochem. 1988; 176: 21-30Google Scholar) and PrPSc(23Turk E. Teplow D.B. Hood L.E. Prusiner S.B. Eur. J. Biochem. 1988; 176: 21-30Google Scholar, 24Hope J. Morton L.J.D. Farquhar C.F. Multhaup G. Beyreuther K. Kimberlin R.H. EMBO J. 1986; 5: 2591-2597Google Scholar, 25Bolton D.C. Bendheim P.E. Marmorstein A.D. Potempska A. Arch. Biochem. Biophys. 1987; 258: 579-590Google Scholar) revealed the posttranslational loss of an N-terminal 22-residue signal peptide. A 23-residue C-terminal peptide is also lost upon addition of a glycosylphosphatidylinositol (GPI) anchor, identified by the release of ~2.8 mol of ethanolamine/mol of PrP together with myoinositol, phosphate, and stearic acid in acid hydrolysates of PrP 27-30(26Stahl N. Borchelt D.R. Hsiao K. Prusiner S.B. Cell. 1987; 51: 229-240Google Scholar). Although PrP has a highly conserved hydrophobic domain, it is not an integral membrane protein but is attached to cell surface membranes by its GPI, which is unusual in that it is sialylated(27Stahl N. Baldwin M.A. Hecker R. Pan K.-M. Burlingame A.L. Prusiner S.B. Biochemistry. 1992; 31: 5043-5053Google Scholar). The majority of PrP molecules carry bi-, tri-, and tetraantennary neutral and N-linked sialylated oligosaccharides at two sites(28Endo T. Groth D. Prusiner S.B. Kobata A. Biochemistry. 1989; 28: 8380-8388Google Scholar, 29Haraguchi T. Fisher S. Olofsson S. Endo T. Groth D. Tarantino A. Borchelt D.R. Teplow D. Hood L. Burlingame A. Lycke E. Kobata A. Prusiner S.B. Arch. Biochem. Biophys. 1989; 274: 1-13Google Scholar). A disulfide bond links the only two cysteines in the mature protein(23Turk E. Teplow D.B. Hood L.E. Prusiner S.B. Eur. J. Biochem. 1988; 176: 21-30Google Scholar). Peptide mapping of PrPSc confirmed that the amino acid sequence corresponds to that predicted from the gene sequence(30Stahl N. Baldwin M.A. Teplow D.B. Hood L. Gibson B.W. Burlingame A.L. Prusiner S.B. Biochemistry. 1993; 32: 1991-2002Google Scholar). The major posttranslational modifications summarized in Fig. 1A are essentially the same for both PrPC and PrPSc, although detailed comparisons of the heterogeneous N-linked sugars and GPI anchors are not completed. PrPC can be solubilized with low concentrations of detergents such as Zwittergent 3-12; it has been purified from hamster and mouse brains by a number of methods, most recently employing detergent solubilization, centrifugation, metal ion affinity chromatography, ion exchange, and lectin affinity(31Pan K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. Huang Z. Fletterick R.J. Cohen F.E. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10962-10966Google Scholar). Purified PrPC does not display the resistance to proteolysis that characterizes PrPSc. Non-denatured PrPSc is insoluble but does not form ordered arrays, whereas PrP 27-30 polymerizes into highly ordered macroscopic amyloid structures(17Prusiner S.B. McKinley M.P. Bowman K.A. Bolton D.C. Bendheim P.E. Groth D.F. Glenner G.G. Cell. 1983; 35: 349-358Google Scholar). Solubilization or disruption of these rod-shaped amyloids by denaturants such as boiling SDS, alkali, or guanidinium thiocyanate inactivate prion infectivity(32Prusiner S.B. Groth D. Serban A. Stahl N. Gabizon R. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 2793-2797Google Scholar). On negatively stained electron micrographs, PrPC and PrPSc appear as amorphous deposits but were immunolabeled with α-PrP antibodies; PrP 27-30 rods were readily visualized without immunolabeling (Fig. 2)(31Pan K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. Huang Z. Fletterick R.J. Cohen F.E. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10962-10966Google Scholar). The polymerization of PrP 27-30 into amyloids and the presence of PrP amyloid plaques in some prion diseases have led to assumptions that amyloid formation is essential for the formation of PrPSc(33Gajdusek D.C. J. Neuroimmunol. 1988; 20: 95-110Google Scholar, 34Gajdusek D.C. Fields B.N. Knipe D.M. Chanock R.M. Hirsch M.S. Melnick J.L. Monath T.P. Roizman B. Virology. Raven Press, New York1990: 2289-2324Google Scholar, 35Gajdusek D.C. Gibbs Jr., C.J. Goldstein A. Biomedical Advances in Aging. Plenum Publishing Corp., New York1990: 3-24Google Scholar, 36Come J.H. Fraser P.E. Lansbury Jr., P.T. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 5959-5963Google Scholar, 37Jarrett J.T. Lansbury Jr., P.T. Cell. 1993; 73: 1055-1058Google Scholar, 38Kocisko D.A. Come J.H. Priola S.A. Chesebro B. Raymond G.J. Lansbury Jr., P.T. Caughey B. Nature. 1994; 370: 471-474Google Scholar). However, PrPSc can be formed in the absence of amyloid, and the presence of amyloid plaques is not obligatory for prion diseases. The existence of different prion isolates or strains with characteristic incubation times and pathology poses an enigma(39Dickinson A.G. Meikle V.M.H. Fraser H. J. Comp. Pathol. 1968; 78: 293-299Google Scholar, 40Fraser H. Dickinson A.G. J. Comp. Pathol. 1973; 83: 29-40Google Scholar, 41Hecker R. Taraboulos A. Scott M. Pan K.-M. Torchia M. Jendroska K. DeArmond S.J. Prusiner S.B. Genes & Dev. 1992; 6: 1213-1228Google Scholar, 42Bruce M. Chree A. McConnell I. Foster J. Pearson G. Fraser H. Philos. Trans. R. Soc. Lond. B Biol. Sci. 1994; 343: 405-411Google Scholar). As prion diseases are apparently caused by an abnormal conformation of PrPSc, different strains might correspond to distinct stable conformations of PrPSc. Alternatively, a second component might mediate the properties of PrPSc. Still another possibility is a labile or infrequent covalent change that induces different properties in PrPSc. The physical properties of PrPSc have made it difficult to obtain for recently has a purification of PrPC been for K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. Huang Z. Fletterick R.J. Cohen F.E. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10962-10966Google Scholar). Fourier transform infrared and have on the of PrP PrPSc, and K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. Huang Z. Fletterick R.J. Cohen F.E. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10962-10966Google Scholar) (Fig. PrPC is essentially and devoid of whereas the of PrPSc is K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. Huang Z. Fletterick R.J. Cohen F.E. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 10962-10966Google Scholar) and the PrP than K.-M. Baldwin M. Nguyen J. Gasset M. Serban A. Groth D. Mehlhorn I. 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Natl. Acad. Sci. U. S. A. 1994; Scholar). PrP have a and form The sequence which and is to in at with G. N. R. E. M. F. Nature. 1993; Scholar, C. L. L. E. L. F. G. F. M. Biochem. Biophys. 1993; Scholar). A of protein for the is and hydrophobic identified the PrP sequence as J.T. Lansbury Jr., P.T. Biochemistry. 1992; 31: formation was in the peptide with and J.H. Fraser P.E. Lansbury Jr., P.T. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 5959-5963Google Scholar). The properties of PrP were to be by the presence of the and P. M. J. B. Gajdusek D.C. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: Scholar). to of PrPSc formation is by a of the known PrP only of which a change to an amino acid a J.M. Nature. 1993; Scholar, Jr., Nature. 1994; Scholar). that the the most with the transmissible disease in Groth D. Scott M. Serban H. D. Foster D. Torchia M. DeArmond S.J. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1994; Scholar). is likely that can be identified that have an on the conformational transition which is that all but of the known within the to PrP 27-30 and are within or to the (Fig. However, of of an acid of the PrP 27-30 also cause prion F. M. R. Collinge J. D. R.M. Hsiao K. Prusiner S.B. Lancet. 1989; 1: Scholar, M. M. R. R.M. T. F. Collinge J. G. J. C. R. Brain. 1992; Scholar, P. D. L. H. Gibbs Gajdusek D.C. Proc. Natl. Acad. Sci. U. S. A. 1991; Scholar, P. B.W. L. K. Gibbs Jr., C.J. Gajdusek D.C. 1993; Scholar). of the to the infectivity is by of the rods in R. McKinley M.P. Groth D.F. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1988; is not a for of by a of a of PrPSc as the E. Groth D.F. Gabizon R. Prusiner S.B. Virology. 1988; Scholar). The for PrPC reveals hydrophobic that be in Z. Baldwin M.A. Fletterick R.J. Prusiner S.B. Cohen F.E. Proc. Natl. Acad. Sci. U. S. 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Diseases of the Press, New Scholar). is to the of PrPC and PrPSc that in their amino acid that both mouse and hamster PrP can be infected with mouse or hamster the that result from these their are resistant to scrapie they are to hamster with hamster H. A. A. P. M. Weissmann C. Cell. 1993; 73: Scholar, S.B. Groth D. Serban A. R. Foster D. Torchia M. D. DeArmond S.J. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: Scholar). can be with the of hamster to is the is hamster than mouse which at two J. Baldwin M.A. Cohen F.E. Prusiner S.B. Biochemistry. Scholar). to PrP sequence mouse and human only of disease with incubation times with human CJD prions. the of was not the human PrP gene was in A PrP gene the human sequence in the from to was into the all of which a fatal illness within of with CJD Scott M. Hsiao Foster D. Torchia M. Collinge J. DeArmond S.J. Prusiner S.B. Proc. Natl. Acad. Sci. U. S. A. 1994; Scholar). that a such as a a in the of PrPC into PrPSc. The Syrian hamster (SHa) and mouse are that can with mouse protein while the affinity of human PrP for protein is The PrPC a for by or other derived from human or in the of of CJD to and not P. Gibbs Jr., C.J. P. D.M. M.P. A. Gajdusek D.C. Ann. Neurol. 1994; 35: Scholar). PrP were to which amino acid were most in H.M. M. L. Cohen F.E. Prusiner S.B. J. Biol. Scholar). with human amino acid from to The most to correspond to the N of PrP 27-30 and the highly are in the whereas two that are for human have a at is that the majority of the sequence the predicted of in to the which within or to these Z. Baldwin M.A. Fletterick R.J. Prusiner S.B. Cohen F.E. Proc. Natl. Acad. Sci. U. S. A. 1994; Scholar). of the of is in the by the of cow disease in R.A. R.F. J.M. J. Gen. Virol. 1994; Scholar). present of prion diseases is to a of using a of cell and protein and of have new of in of of the PrP to be to infectivity by in of or proteins of PrP or a second is for prion an of the the of PrPC to PrPSc the or of to the of prion diseases. prion and disease Borchelt D.R. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: Scholar) common of in these A to prion formation of some in Science. 1994; Scholar). it that the new of prion has than