scholarly journals Copper induces increased beta-sheet content in the scrapie-susceptible ovine prion protein PrPVRQ compared with the resistant allelic variant PrPARR

2004 ◽  
Vol 380 (1) ◽  
pp. 273-282 ◽  
Author(s):  
Edmond WONG ◽  
Alana M. THACKRAY ◽  
Raymond BUJDOSO
Keyword(s):  
Amino Acid ◽  
Prion Protein ◽  
Metal Ion ◽  
Cd Spectroscopy ◽  
Proteinase K ◽  
Copper Binding ◽  
Protein Amino Acid ◽  
Allelic Variants ◽  
Β Sheet

Prion diseases are characterized by conformational change in the copper-binding protein PrP (prion protein). Polymorphisms in ovine PrP at amino acid residues 136, 154 and 171 are associated with variation in susceptibility to scrapie. PrPVRQ [PrP(Val136/Arg154/Gln171)] or PrPARQ [PrP(Ala136/Arg154/Gln171)] animals show susceptibility to scrapie, whereas those that express Ala136/Arg154/Arg171 (PrPARR) show resistance. Results are presented here that show PrPVRQ and PrPARR display different conformational responses to metal-ion interaction. At 37 °C copper induced different levels of β-sheet content in the allelic variants of ovine full-length prion protein (amino acid 25–232). PrPVRQ showed a significant increase in β-sheet content when exposed to copper at 37 °C, whereas PrPARR remained relatively unchanged. The conversion of α-helical PrPVRQ to β-sheet form was shown by CD spectroscopy and the decreased binding of C-terminal specific monoclonal anti-PrP antibodies. This conversion to an increased β-sheet form did not occur with truncated PrPVRQ (amino acids 89–233), which demonstrates that additional metal-binding sites outside of the N-terminus may not overtly influence the overall structure of ovine PrP. Despite the difference in β-sheet content, both the scrapie-susceptible and -resistant allelic forms of ovine PrP acquired resistance to proteinase K digestion following exposure to copper at 37 °C, suggesting the potential for disease-associated PrPARR to accumulate in vivo. Our present study demonstrates that allelic variants of ovine PrP differ in their structure and response to the interaction with copper. These observations will contribute to a better understanding of the mechanism of susceptibility and resistance to prion disease.

scholarly journals Isolation and characterization of a polymerized prion protein

2002 ◽  
Vol 364 (1) ◽  
pp. 81-87 ◽  
Author(s):  
Bao-Yuan LU ◽  
Jui-Yoa CHANG
Keyword(s):  
Prion Protein ◽  
Cd Spectroscopy ◽  
Reversed Phase ◽  
Size Exclusion ◽  
Proteinase K ◽  
Guanidinium Chloride ◽  
Isolation And Characterization ◽  
Size Exclusion Hplc ◽  
Β Sheet

A polymerized form of recombinant mouse prion protein (mPrP) domain 23–231 [mPrP-(23–231)], designated mPrP-z, was generated at acidic pH (pH 2–5) in the presence of selected concentrations of denaturant (2M guanidinium chloride or 5M urea). This isoform of mPrP is stable in acidic solution after removal of denaturant. It can be isolated and purified using reversed-phase HPLC or size-exclusion HPLC. mPrP-z bears structural properties that partially resemble those of scrapie prion. Unlike the native mPrP-(23–231) (mPrP-N), mPrP-z exhibits a high content of β-sheet structure, as shown by CD spectroscopy, and exists as an oligomer with an approximate molecular mass of 340000Da, as measured by light scattering. However, similarly to mPrP-N, mPrP-z contains the intact disulphide bond and is sensitive to digestion by proteinase K.

scholarly journals Compositional Determinants of Prion Formation in Yeast

2009 ◽  
Vol 30 (1) ◽  
pp. 319-332 ◽  
Author(s):  
James A. Toombs ◽  
Blake R. McCarty ◽  
Eric D. Ross
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Amyloid Formation ◽  
Yeast Prion ◽  
Hydrophobic Residues ◽  
Predictive Methods ◽  
Infectious Proteins ◽  
Β Sheet ◽  
Eukaryotic Genomes

ABSTRACT Numerous prions (infectious proteins) have been identified in yeast that result from the conversion of soluble proteins into β-sheet-rich amyloid-like protein aggregates. Yeast prion formation is driven primarily by amino acid composition. However, yeast prion domains are generally lacking in the bulky hydrophobic residues most strongly associated with amyloid formation and are instead enriched in glutamines and asparagines. Glutamine/asparagine-rich domains are thought to be involved in both disease-related and beneficial amyloid formation. These domains are overrepresented in eukaryotic genomes, but predictive methods have not yet been developed to efficiently distinguish between prion and nonprion glutamine/asparagine-rich domains. We have developed a novel in vivo assay to quantitatively assess how composition affects prion formation. Using our results, we have defined the compositional features that promote prion formation, allowing us to accurately distinguish between glutamine/asparagine-rich domains that can form prion-like aggregates and those that cannot. Additionally, our results explain why traditional amyloid prediction algorithms fail to accurately predict amyloid formation by the glutamine/asparagine-rich yeast prion domains.

scholarly journals Efficient Conversion of Normal Prion Protein (PrP) by Abnormal Hamster PrP Is Determined by Homology at Amino Acid Residue 155

2001 ◽  
Vol 75 (10) ◽  
pp. 4673-4680 ◽  
Author(s):  
Suzette A. Priola ◽  
Joëlle Chabry ◽  
Kaman Chan
Keyword(s):  
Amino Acid ◽  
Prion Protein ◽  
Amino Acid Residue ◽  
Animal Species ◽  
Alpha Helix ◽  
Proteinase K ◽  
Transmissible Spongiform Encephalopathies ◽  
Spongiform Encephalopathies ◽  
A Cell ◽  
Resistant Product

ABSTRACT In the transmissible spongiform encephalopathies, disease is closely associated with the conversion of the normal proteinase K-sensitive host prion protein (PrP-sen) to the abnormal proteinase K-resistant form (PrP-res). Amino acid sequence homology between PrP-res and PrP-sen is important in the formation of new PrP-res and thus in the efficient transmission of infectivity across species barriers. It was previously shown that the generation of mouse PrP-res was strongly influenced by homology between PrP-sen and PrP-res at amino acid residue 138, a residue located in a region of loop structure common to PrP molecules from many different species. In order to determine if homology at residue 138 also affected the formation of PrP-res in a different animal species, we assayed the ability of hamster PrP-res to convert a panel of recombinant PrP-sen molecules to protease-resistant PrP in a cell-free conversion system. Homology at amino acid residue 138 was not critical for the formation of protease-resistant hamster PrP. Rather, homology between PrP-sen and hamster PrP-res at amino acid residue 155 determined the efficiency of formation of a protease-resistant product induced by hamster PrP-res. Structurally, residue 155 resides in a turn at the end of the first alpha helix in hamster PrP-sen; this feature is not present in mouse PrP-sen. Thus, our data suggest that PrP-res molecules isolated from scrapie-infected brains of different animal species have different PrP-sen structural requirements for the efficient formation of protease-resistant PrP.

scholarly journals The Prion Protein Octarepeat Domain Forms Transient β-sheet Structures Upon Residue-Specific Cu(II) and Zn(II) Binding

2021 ◽  
Author(s):  
Maciej Gielnik ◽  
Aneta Szymanska ◽  
Xiaolin Dong ◽  
Jyri Jarvet ◽  
Zeljko M. Svedruzic ◽  
...  
Keyword(s):  
Metal Ions ◽  
Prion Protein ◽  
Metal Binding ◽  
Cd Spectroscopy ◽  
Cellular Prion Protein ◽  
Transmissible Spongiform Encephalopathies ◽  
Amyloid Formation ◽  
Spectroscopy Data ◽  
Spongiform Encephalopathies ◽  
Β Sheet

Misfolding of the cellular prion protein (PrPC) is associated with the development of fatal neurodegenerative diseases called transmissible spongiform encephalopathies (TSEs). Metal ions appear to play a crucial role in the protein misfolding, and metal imbalance may be part of TSE pathologies. PrPC is a combined Cu(II) and Zn(II) metal binding protein, where the main metal binding site is located in the octarepeat (OR) region. Here, we used biophysical methods to characterize Cu(II) and Zn(II) binding to the isolated OR region. Circular dichroism (CD) spectroscopy data suggest that the OR domain binds up to four Cu(II) ions or two Zn(II) ions. Upon metal binding, the OR region seems to adopt a transient antiparallel β-sheet hairpin structure. Fluorescence spectroscopy data indicates that under neutral conditions, the OR region can bind both Cu(II) and Zn(II) ions, whereas under acidic conditions it binds only Cu(II) ions. Molecular dynamics simulations suggest that binding of both metal ions to the OR region results in formation of β-hairpin structures. As formation of β-sheet structures is a first step towards amyloid formation, we propose that high concentrations of either Cu(II) or Zn(II) ions may have a pro-amyloid effect in TSEs.

scholarly journals Synergistic and strain-specific effects of bovine spongiform encephalopathy and scrapie prions in the cell-free conversion of recombinant prion protein

2006 ◽  
Vol 87 (12) ◽  
pp. 3753-3761 ◽  
Author(s):  
Martin Eiden ◽  
Gottfried J. Palm ◽  
Winfried Hinrichs ◽  
Ulrich Matthey ◽  
Ralph Zahn ◽  
...  
Keyword(s):  
Bovine Spongiform Encephalopathy ◽  
Prion Protein ◽  
De Novo ◽  
Proteinase K ◽  
Spongiform Encephalopathy ◽  
Cooperative Action ◽  
Specific Effects ◽  
Scrapie Strains ◽  
Recombinant Prion Protein

This study describes the conversion of murine PrPC by PrPSc from three different mouse scrapie strains (ME7, 87V and 22A) and from a mouse-passaged bovine spongiform encephalopathy (BSE) strain (BSE/Bl6). This was demonstrated by a modified, non-radioactive, cell-free conversion assay using bacterial prion protein, which was converted into a proteinase K (PK)-resistant fragment designated PrPres. Using this assay, newly formed PrPres could be detected by an antibody that discriminated de novo PrPres and the original PrPSc seed. The results suggested that PrPres formation occurs in three phases: the first 48 h when PrPres formation is delayed, followed by a period of substantially accelerated PrPres formation and a plateau phase when a maximum concentration of PrPres is reached after 72 h. The conversion of prokaryotically expressed PrPC by ME7 and BSE prions led to unglycosylated, PK-digested, abnormal PrPres fragments, which differed in molecular mass by 1 kDa. Therefore, prion strain phenotypes were retained in the cell-free conversion, even when recombinant PrPC was used as the substrate. Moreover, co-incubation of ME7 and BSE prions resulted in equal amounts of both ME7- and BSE-derived PrPres fragments (as distinguished by their different molecular sizes) and also in a significantly increased total amount of de novo-generated PrPres. This was found to be more than twice the amount of either strain when incubated separately. This result indicates a synergistic effect of both strains during cell-free conversion. It is not yet known whether such a cooperative action between BSE and scrapie prions also occurs in vivo.

Correlation of the β-sheet crystal size in silk fibers with the protein amino acid sequence

Soft Matter ◽  
2007 ◽  
Vol 3 (7) ◽  
pp. 877-882 ◽  
Author(s):  
Lawrence F. Drummy ◽  
B. L. Farmer ◽  
Rajesh R. Naik
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Crystal Size ◽  
Protein Amino Acid ◽  
Silk Fibers ◽  
Protein Amino Acid Sequence ◽  
Β Sheet

scholarly journals Structural and Physical Properties of a Necrosis-Inducing Toxin from Pyrenophora tritici-repentis

1997 ◽  
Vol 87 (2) ◽  
pp. 154-160 ◽  
Author(s):  
Hui-Fen Zhang ◽  
Leonard J. Francl ◽  
James G. Jordahl ◽  
Steven W. Meinhardt
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Structure Prediction ◽  
Secondary Structure Prediction ◽  
Cd Spectroscopy ◽  
Tan Spot ◽  
Pyrenophora Tritici Repentis ◽  
Membrane Adhesion ◽  
Α Helix ◽  
Β Sheet

Cultivar-specific toxic metabolites of Pyrenophora tritici-repentis are involved in the appearance of necrotic and chlorotic foliar lesions characteristic of tan spot. A P. tritici-repentis necrosis-inducing toxin, Ptr necrosis toxin, was purified from isolate 86-124, sequenced by gas-phase amino acid microsequencing, and characterized by circular dichroism (CD) spectroscopy and isoelectric focusing. The purified protein had a similar amino acid composition and molecular weight as previously reported. Analysis of the CD spectrum from 178 to 250 nm indicated a protein consisting of 13% α-helix, 36% antiparallel β-sheet, 25% turns, and 25% other structures. The Ptr necrosis toxin from isolate 86-124 has an isoelectric point near pH 10. Using overlapping proteolytic fragments obtained from the toxin, a sequence of 101 continuous amino acids was obtained, but the amino terminus was blocked and 9 to 16 amino acids could not be sequenced. Secondary structure prediction based on the amino acid sequence indicated a β-sheet protein with little α-helix, which is in agreement with the structure determined by CD spectroscopy. Sequence analysis indicated the presence of a possible membrane adhesion site and several possible phosphorylation sites that may be involved in phytotoxicity.

scholarly journals A reassessment of copper(II) binding in the full-length prion protein

2006 ◽  
Vol 399 (3) ◽  
pp. 435-444 ◽  
Author(s):  
Mark A. Wells ◽  
Graham S. Jackson ◽  
Samantha Jones ◽  
Laszlo L. P. Hosszu ◽  
C. Jeremy Craven ◽  
...  
Keyword(s):  
Prion Protein ◽  
Metal Ion ◽  
Tandem Repeats ◽  
Binding Mode ◽  
Full Length ◽  
Alternative Mode ◽  
Histidine Residues ◽  
Terminal Domain

It has been shown previously that the unfolded N-terminal domain of the prion protein can bind up to six Cu2+ ions in vitro. This domain contains four tandem repeats of the octapeptide sequence PHGGGWGQ, which, alongside the two histidine residues at positions 96 and 111, contribute to its Cu2+ binding properties. At the maximum metal-ion occupancy each Cu2+ is co-ordinated by a single imidazole and deprotonated backbone amide groups. However two recent studies of peptides representing the octapeptide repeat region of the protein have shown, that at low Cu2+ availability, an alternative mode of co-ordination occurs where the metal ion is bound by multiple histidine imidazole groups. Both modes of binding are readily populated at pH 7.4, while mild acidification to pH 5.5 selects in favour of the low occupancy, multiple imidazole binding mode. We have used NMR to resolve how Cu2+ binds to the full-length prion protein under mildly acidic conditions where multiple histidine co-ordination is dominant. We show that at pH 5.5 the protein binds two Cu2+ ions, and that all six histidine residues of the unfolded N-terminal domain and the N-terminal amine act as ligands. These two sites are of sufficient affinity to be maintained in the presence of millimolar concentrations of competing exogenous histidine. A previously unknown interaction between the N-terminal domain and a site on the C-terminal domain becomes apparent when the protein is loaded with Cu2+. Furthermore, the data reveal that sub-stoichiometric quantities of Cu2+ will cause self-association of the prion protein in vitro, suggesting that Cu2+ may play a role in controlling oligomerization in vivo.

scholarly journals Specific Amyloid Binding of Polybasic Peptides In Vivo Is Retained by β-Sheet Conformers but Lost in the Disrupted Coil and All D-Amino Acid Variants

2017 ◽  
Vol 19 (5) ◽  
pp. 714-722 ◽  
Author(s):  
Jonathan S. Wall ◽  
Angela Williams ◽  
Tina Richey ◽  
Alan Stuckey ◽  
Craig Wooliver ◽  
...  
Keyword(s):  
Amino Acid ◽  
Β Sheet ◽  
Amino Acid Variants

scholarly journals Prion Protein Amino Acid Determinants of Differential Susceptibility and Molecular Feature of Prion Strains in Mice and Voles

2008 ◽  
Vol 4 (7) ◽  
pp. e1000113 ◽  
Author(s):  
Umberto Agrimi ◽  
Romolo Nonno ◽  
Giacomo Dell'Omo ◽  
Michele Angelo Di Bari ◽  
Michela Conte ◽  
...  
Keyword(s):  
Amino Acid ◽  
Prion Protein ◽  
Differential Susceptibility ◽  
Protein Amino Acid ◽  
Molecular Feature ◽  
Prion Strains
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