Small, Highly Structured RNAs Participate in the Conversion of Human Recombinant PrPSen to PrPRes in Vitro

Adler, Victor; Zeiler, Brian; Kryukov, V. A.; Kascsak, Richard J.; Rubenstein, Richard; Grossman, Abraham

doi:10.1016/s0022-2836(03)00919-7

Cited by 101 publications

(110 citation statements)

References 35 publications

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“…1 Synthetic nucleic acids, in solution and in vitro, can catalyze conversion of recombinant and cellular PrP C to PrP Sc as evidenced from secondary structural studies of the protein and proteinase K (PK) resistance properties. [4][5][6][7] Our studies also indicated that bending and unwinding of nucleic acid occurred by prion protein and natural poly amines (spermine and spermidine) can inhibit this interaction. 8,9 Highly structured small RNA binds to PrP C at neutral pH and yields Proteinase K resistant component in the presence of cellular cofactors.…”

Section: Introductionsupporting

confidence: 54%

“…8,9 Highly structured small RNA binds to PrP C at neutral pH and yields Proteinase K resistant component in the presence of cellular cofactors. 6 Studies using brain tissues have shown that an endogenous 300 nucleotide long RNA (100 kDa) can convert endogenous PrP C to Proteinase K resistant form in vitro. 7,10,11 These results indicate that nucleic acid can act as a cofactor for the conversion of PrP C to PrP Sc and can be the active transmissible spongiform encephalopathy (TSE) agent.…”

Section: Introductionmentioning

confidence: 99%

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Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent

Bera

Nandi

2014

Protein Science

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Nucleic acid can catalyze the conversion of a-helical cellular prion protein to b-sheet rich Proteinase K resistant prion protein oligomers and amyloid polymers in vitro and in solution. Because unfolding of a protein molecule from its ordered a-helical structure is considered to be a necessary step for the structural conversion to its b-sheet rich isoform, we have studied the unfolding of the a-helical globular 121-231 fragment of mouse recombinant prion protein in the presence of different nucleic acids at neutral and acid pH. Nucleic acids, either single or double stranded, do not have any significant effect on the secondary structure of the protein fragment at neutral pH; however the protein secondary structure is modified by the nucleic acids at pH 5. Nucleic acids do not show any significant effect on the temperature induced unfolding of the globular prion protein domain at neutral pH which, however, undergoes a gross conformational change at pH 5 as evidenced from the lowering of the midpoint of thermal denaturation temperatures, Tm, of the protein. The extent of Tm decrease shows a dependence on the nature of nucleic acid. The interaction of nucleic acid with the nonpolar groups exposed from the protein interior at pH 5 probably contributes substantially to the unfolding process of the protein.

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Section: Introductionsupporting

confidence: 54%

Section: Introductionmentioning

confidence: 99%

Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent

Bera

Nandi

2014

Protein Science

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“…Further studies are required to distinguish between these possibilities and to identify the specific endogenous polyanions that facilitate prion propagation in vivo. Several classes of negatively charged macromolecules could potentially serve as cofactors for prion propagation, including: nucleic acids (17,(32)(33)(34)(35)(36)(37)(38), glycosaminoglycans (14-16), phospholipid-rich membranes (39)(40)(41)(42), and chaperone proteins (43). Interestingly, it has been proposed that polyanions could play a broad role in protein folding and misfolding, and the ability of polyanions to facilitate prion conversion may represent a specific example of that general concept (44).…”

Section: Discussionmentioning

confidence: 99%

Formation of native prions from minimal components in vitro

Deleault¹,

Harris²,

Rees³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

585

635

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The conformational change of a host protein, PrP C , into a disease-associated isoform, PrP Sc , appears to play a critical role in the pathogenesis of prion diseases such as Creutzfeldt–Jakob disease and scrapie. However, the fundamental mechanism by which infectious prions are produced in neurons remains unknown. To investigate the mechanism of prion formation biochemically, we conducted a series of experiments using the protein misfolding cyclic amplification (PMCA) technique with a preparation containing only native PrP C and copurified lipid molecules. These experiments showed that successful PMCA propagation of PrP Sc molecules in a purified system requires accessory polyanion molecules. In addition, we found that PrP Sc molecules could be formed de novo from these defined components in the absence of preexisting prions. Inoculation of samples containing either prion-seeded or spontaneously generated PrP Sc molecules into hamsters caused scrapie, which was transmissible on second passage. These results show that prions able to infect wild-type hamsters can be formed from a minimal set of components including native PrP C molecules, copurified lipid molecules, and a synthetic polyanion.

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“…forms of Prnp,"⌬PrP") have traditionally been equated with protein activities (24,25,72), because unusual secondary structures have been proposed for Prnp mRNA (73)(74)(75) and conversion of PrP C to protease-resistant forms is facilitated by cellular RNAs (76,77), we also considered RNA-based mechanisms. To exclude biological properties of this ribonucleotide sequence as a basis for pro-apoptotic activity, we created a Prnd allele with tandem stop codons inserted just upstream of this position (Dpl91,92terter).…”

Section: Activity Determinants In Prp C and Dplmentioning

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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Small, Highly Structured RNAs Participate in the Conversion of Human Recombinant PrPSen to PrPRes in Vitro

Cited by 101 publications

References 35 publications

Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent

Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent

Formation of native prions from minimal components in vitro

Genetic Mapping of Activity Determinants within Cellular Prion Proteins

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