Mass analysis by scanning transmission electron microscopy and electron diffraction validate predictions of stacked beta-solenoid model of HET-s prion fibrils.
Fungal prions are infectious filamentous polymers of proteins that are soluble in uninfected cells. In its prion form, the HET-s protein of Podospora anserina participates in a fungal self/non-self recognition phenomenon called heterokaryon incompatibility. Like other prion proteins, HET-s has a so-called "prion domain" (its C-terminal region, HET-s-(218-289)) that is responsible for induction and propagation of the prion in vivo and for fibril formation in vitro. Prion fibrils are thought to have amyloid backbones of polymerized prion domains. A relatively detailed model has been proposed for prion domain fibrils of HET-s based on a variety of experimental constraints (Ritter, C., Maddelein, M. L., Siemer, A. B., Luhrs, T., Ernst, M., Meier, B. H., Saupe, S. J., and Riek, R. (2005) Nature 435, 844-848). To test specific predictions of this model, which envisages axial stacking of beta-solenoids with two coils per subunit, we examined fibrils by electron microscopy. Electron diffraction gave a prominent meridional reflection at (0.47 nm)(-1), indicative of cross-beta structure, as predicted. STEM (scanning transmission electron microscopy) mass-per-unit-length measurements yielded 1.02 +/- 0.16 subunits per 0.94 nm, in agreement with the model prediction (1 subunit per 0.94 nm). This is half the packing density of approximately 1 subunit per 0.47 nm previously obtained for fibrils of the yeast prion proteins, Ure2p and Sup35p, whence it follows that the respective amyloid architectures are basically different.
Sen
Anindito
A
Laboratory of Structural Biology, National Institute of Arthritis, Musculoskeletal, and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Baxa
Ulrich
U
Simon
Martha N
MN
Wall
Joseph S
JS
Sabate
Raimon
R
Saupe
Sven J
SJ
Steven
Alasdair C
AC
D000682:Amyloid / Q000737:chemistry / Q000648:ultrastructure*; D005656:Fungal Proteins / Q000737:chemistry*; D017348:Microscopy, Electron, Scanning Transmission / Q000379:methods; D008958:Models, Molecular; D048468:Podospora / Q000737:chemistry*; D011328:Prions / Q000737:chemistry*; D017434:Protein Structure, Tertiary
D000682:Amyloid; D005656:Fungal Proteins; C083891:HET-S protein, Podospora anserina; D011328:Prions