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Published on: July 16, 2008
Prion Amyloid Polymorphs - The Tag Might Change It All
Luc Bousset1, Nina Luckgei2, Mehdi Kabani1
1Institut Francois Jacob (MIRCen), CEA and Laboratory of Neurodegenerative Diseases, CNRS, Fontenay-aux-Roses, France.
Abstract:
Sup35p is a protein from Saccharomyces cerevisiae. It can propagate using a prion-like mechanism, which means that it can recruit non-prion soluble Sup35p into insoluble fibrils. Sup35p is a large protein showing three distinct domains, N, M and an extended globular domain. We have previously studied the conformations of the full-length and truncated NM versions carrying poly-histidine tags on the N-terminus. Comparison with structural data from C-terminally poly-histidine tagged NM from the literature surprisingly revealed discrepancies. Here we investigated fibrils from the untagged, as well as a C-terminally poly-histidine tagged NM construct, using solid-state NMR. We find that the conformation of untagged NM is very close to the N-terminally tagged NM and confirms our previous findings. The C-terminal poly-histidine tag, in contrast, drastically changes the NM fibril structure, and yields data consistent with results obtained previously on this construct. We conclude that the C-terminally located Sup35p globular domain influences the structure of the fibrillar core at the N domain, as previously shown. We further conclude, based on the present data, that small tags on NM C-terminus have a substantial, despite different, impact. Modifications at this remote localization thus shows an unexpected influence on the fibril structure, and importantly also its propensity to induce [PSI+].
Insights
Polyhistidine tags on Sup35p NM fibrils significantly alter their structure. C-terminal tags dramatically change fibril conformation, impacting [PSI+] induction, unlike untagged or N-terminally tagged versions.
Area of Science:
- Biochemistry
- Structural Biology
- Molecular Biology
Background:
- Sup35p protein from Saccharomyces cerevisiae propagates via a prion-like mechanism, forming insoluble fibrils from soluble precursors.
- Sup35p comprises N, M, and globular domains; previous studies on tagged NM constructs showed discrepancies in fibril conformation.
Purpose of the Study:
- To investigate the structural impact of C-terminal polyhistidine tags on Sup35p NM fibrils using solid-state NMR.
- To compare fibril conformations of untagged, N-terminally tagged, and C-terminally tagged Sup35p NM constructs.
Main Methods:
- Solid-state Nuclear Magnetic Resonance (NMR) spectroscopy was employed.
- Fibrils from untagged and C-terminally polyhistidine-tagged Sup35p NM constructs were analyzed.
Main Results:
- The conformation of untagged Sup35p NM fibrils closely matched N-terminally tagged versions, validating prior findings.
- A C-terminal polyhistidine tag drastically altered the NM fibril structure, consistent with previous literature on this construct.
- Modifications at the C-terminus, even small tags, exerted a substantial and unexpected influence on fibril structure and [PSI+] induction propensity.
Conclusions:
- The C-terminally located globular domain of Sup35p influences the N-domain fibrillar core structure.
- C-terminal tags on Sup35p NM significantly impact fibril structure and the protein's ability to induce the [PSI+] prion state.
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