Molecular cloning and complete sequence of prion protein cDNA from mouse brain infected with the scrapie agent.

Locht, Camille; Chesebro, Bruce; Race, Richard E.; Keith, Jerry M.

doi:10.1073/pnas.83.17.6372

Cited by 159 publications

(87 citation statements)

References 44 publications

(52 reference statements)

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“…PrP33-35, PrP26-29 and PrP23 -25 (not shown) gave a major N-terminal sequence of Lys-Lys-Arg-Pro-Lys-Pro-Gly-Gly-. This is exactly that predicted by cDNA cloning and on theoretical grounds for the amino-terminal region of the normal hamster PrP protein [lo, 141; this sequence and a similar putative signal sequence have been inferred from the cDNA sequence of mouse PrP mRNA [17]. PrP26-29 and PrP23 -25 also contained minor sequences which began Ser49'57-Trp-Gly-Gln-Pro-His-GlyGly, identifying some components with amino-terminal sequences in the octapeptide repeat segment of the protein.…”

Section: Experiments 2 Andmentioning

confidence: 71%

“…Amino-acid sequencing of PrP proteins in mouse SAF The cycle number refers to the number of residues determined. Major and minor sequences refer to the first amino acids of the most abundant protein sequence or minor sequences, respectively; they are numbered with reference to the PrP sequence inferred from the cDNA copy of mouse PrP [17] (see Fig. 4).…”

Section: Experiments 2 Andmentioning

confidence: 99%

See 1 more Smart Citation

Molecular pathology of scrapie‐associated fibril protein (PrP) in mouse brain affected by the ME7 strain of scrapie

Hope

Multhaup

Reekie

et al. 1988

European Journal of Biochemistry

124

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Scrapie‐associated fibrils (SAF) are disease‐specific structures found in extracts of the brains of animals affected with scrapie. These structures are pathological aggregates of a normal host protein (PrP). Abnormal post‐translational modification of PrP has been suggested to explain its aberrant properties in scrapie‐affected brains and although there is a form of PrP in SAF indistinguishable in size from the protein in uninfected brain, lower‐molecular‐mass variants of PrP are also found in SAF fractions. We report the characterisation of the multiple forms of PrP found in SAF fractions purified from mouse brain affected by the ME7 strain of scrapie. The quantitively major forms of PrP in SAF prepared without the use of proteinase K have the amino‐terminal sequence Lys‐Lys‐Arg‐Pro‐Lys‐Pro‐Gly‐Gly‐, identical to that predicted for the amino‐terminus of normal mouse brain PrP. However N‐terminal cleavage of some PrP does occur in vivo within a domain of repetitive sequences at sites similar to but distinct from those cut by proteinase K in vitro. This suggests the conformation of the protein in aggregates in vivo does not differ extensively from that in detergent‐treated SAF in vitro. We conclude that the size diversity of PrP in SAF is only partly due to N‐terminal proteolysis and is independent of the proteolysis that occurs if proteinase K is used in the purification of SAF. Apart from proteolytic changes in the structure of PrP, we found a novel, as yet unidentified, amino‐acid derivative of the arginine residue at position 3 in mouse PrP, which may predispose PrP to form SAF.

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Section: Experiments 2 Andmentioning

confidence: 71%

Section: Experiments 2 Andmentioning

confidence: 99%

Molecular pathology of scrapie‐associated fibril protein (PrP) in mouse brain affected by the ME7 strain of scrapie

Hope

Multhaup

Reekie

et al. 1988

European Journal of Biochemistry

124

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“…The 13.3-kDa species corresponds to two approximately equimolar species, consistent with peptides commencing at residues 116 and 118 of mouse PrP (44). The cleavage sites between residues 1152116 and 1172118 are not identical to those observed when MoPrP23-231 is purified in the absence of protease inhibitors (cleavage at 1162117, 1182119, and 1202121) (33).…”

Section: Cu(ii)-induced Formation Of Proteinase K-resistant Prp Fragmmentioning

confidence: 84%

Copper(II)-induced Conformational Changes and Protease Resistance in Recombinant and Cellular PrP

Qin¹,

Yang²,

Yang³

et al. 2000

Journal of Biological Chemistry

151

128

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While PrPC rearranges in the area of codons 104 -113 to form PrP Sc during prion infections, the events that initiate sporadic Creutzfeldt-Jakob disease are undefined. As Cu(II) is a putative ligand for PrP C and has been implicated in the pathogenesis of Creutzfeldt-Jakob disease and other neurodegenerative diseases, we investigated the structural effects of binding. Incubation of brain microsomes with Cu(II) generated ϳ30-kDa proteinase K-resistant PrP. Cu(II) had little effect on fresh recombinant PrP23-231, but aged protein characterized by conversion of Asn-107 to Asp decreased ␣-helical content by ϳ30%, increased ␤-sheet content 100%, formed aggregates, and acquired proteinase K resistance in the presence of Cu(II). These transitions took place without need for acid pH, organic solvents, denaturants, or reducing agents. Since conversion of Asn to Asp proceeds by a spontaneous pathway involving deamidation, our data suggest that covalent variants of PrP C arising in this manner may, in concert with Cu(II), generate PrP Sc -like species capable of initiating sporadic prion disease.

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“…The position of the resulting 5-amino acid insertion was established by sequencing. The wt Prnp allele used as a basis for mutagenesis was derived from a Prnp a cDNA (32). This was modified to include a NotI site in the 5Ј leader sequence encoded by exon 2 (33) adjacent to a naturally occurring KpnI site and a second NotI site 80 bp downstream of the termination codon.…”

Section: Methodsmentioning

confidence: 99%

Genetic Mapping of Activity Determinants within Cellular Prion Proteins

Drisaldi

Coomaraswamy

Mastrangelo

et al. 2004

Journal of Biological Chemistry

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The PrP-like Doppel (Dpl) protein causes apoptotic death of cerebellar neurons in transgenic mice, a process prevented by expression of the wild type (wt) cellular prion protein, PrP C . Internally deleted forms of PrP C resembling Dpl such as PrP⌬32-121 produce a similar PrP C -sensitive pro-apoptotic phenotype in transgenic mice. Here we demonstrate that these phenotypic attributes of wt Dpl, wt PrP C , and PrP⌬132-121 can be accurately recapitulated by transfected mouse cerebellar granule cell cultures. This system was then explored by mutagenesis of the co-expressed prion proteins to reveal functional determinants. By this means, neuroprotective activity of wt PrP C was shown to be nullified by a deletion of the N-terminal charged region implicated in endocytosis and retrograde axonal transport (PrP⌬23-28), by deletion of all five octarepeats (PrP⌬51-90), or by glycine replacement of four octarepeat histidine residues required for selective binding of copper ions (Prnp"H/G"). In the case of Dpl, overlapping deletions defined a requirement for the gene interval encoding helices B and B (Dpl⌬101-125). These data suggest contributions of copper binding and neuronal trafficking to wt PrP C function in vivo and place constraints upon current hypotheses to explain Dpl/PrP C antagonism by competitive ligand binding. Further implementation of this assay should provide a fuller understanding of the attributes and subcellular localizations required for activity of these enigmatic proteins.

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Molecular cloning and complete sequence of prion protein cDNA from mouse brain infected with the scrapie agent.

Cited by 159 publications

References 44 publications

Molecular pathology of scrapie‐associated fibril protein (PrP) in mouse brain affected by the ME7 strain of scrapie

Molecular pathology of scrapie‐associated fibril protein (PrP) in mouse brain affected by the ME7 strain of scrapie

Copper(II)-induced Conformational Changes and Protease Resistance in Recombinant and Cellular PrP

Genetic Mapping of Activity Determinants within Cellular Prion Proteins

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