RT Journal Article
T1 Structural organization of mammalian prions as probed by limited proteolysis
A1 Vázquez Fernández, Ester
A1 Alonso Lorenzo, Jana
A1 Pastrana González, Miguel Ángel
A1 Ramos Amigo, Adriana
A1 Stitz, Lothar
A1 Vidal, Enric
A1 Dynin, Irina
A1 Petsch, Benjamin
A1 Silva, Christopher J.
A1 Rodríguez Requena, Jesús
AB Elucidation of the structure of PrPSc continues to be one major challenge in prion research. The mechanism of propagation of these infectious agents will not be understood until their structure is solved. Given that high resolution techniques such as NMR or X-ray crystallography cannot be used, a number of lower resolution analytical approaches have been attempted. Thus, limited proteolysis has been successfully used to pinpoint flexible regions within prion multimers (PrPSc). However, the presence of covalently attached sugar antennae and glycosylphosphatidylinositol (GPI) moieties makes mass spectrometry-based analysis impractical. In order to surmount these difficulties we analyzed PrPSc from transgenic mice expressing prion protein (PrP) lacking the GPI membrane anchor. Such animals produce prions that are devoid of the GPI anchor and sugar antennae, and, thereby, permit the detection and location of flexible, proteinase K (PK) susceptible regions by Western blot and mass spectrometry-based analysis. GPI-less PrPSc samples were digested with PK. PK-resistant peptides were identified, and found to correspond to molecules cleaved at positions 81, 85, 89, 116, 118, 133, 134, 141, 152, 153, 162, 169 and 179. The first 10 peptides (to position 153), match very well with PK cleavage sites we previously identified in wild type PrPSc. These results reinforce the hypothesis that the structure of PrPSc consists of a series of highly PK-resistant β-sheet strands connected by short flexible PK-sensitive loops and turns. A sizeable C-terminal stretch of PrPSc is highly resistant to PK and therefore perhaps also contains β-sheet secondary structure.
PB PLOS
YR 2012
FD 2012
LK http://hdl.handle.net/10347/22019
UL http://hdl.handle.net/10347/22019
LA eng
NO Vázquez-Fernández E, Alonso J, Pastrana MA, Ramos A, Stitz L, Vidal E, et al. (2012) Structural Organization of Mammalian Prions as Probed by Limited Proteolysis. PLoS ONE 7(11): e50111. https://doi.org/10.1371/journal.pone.0050111
NO This work was supported by EC grant FP7 222887 “Priority”
DS Minerva
RD 28 abr 2026