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Orb2 shares common features with the pathological amyloidogenic pathway.

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posted on 28.01.2016 by Rubén Hervás, Liying Li, Amitabha Majumdar, María del Carmen Fernández-Ramírez, Jay R. Unruh, Brian D. Slaughter, Albert Galera-Prat, Elena Santana, Mari Suzuki, Yoshitaka Nagai, Marta Bruix, Sergio Casas-Tintó, Margarita Menéndez, Douglas V. Laurents, Kausik Si, Mariano Carrión-Vázquez

(A) ΔLc and F SMFS histograms of pFS-2 polyproteins carrying the Q/N-rich PLD of Orb2A show a broad mechanical polymorphism in terms of the increased contour length (ΔLc, top) and mechanical stability (bottom), ranging from NM conformers (orange bars) to different M conformers (red bars, n = 106), similar to that found in pathological amyloids [41]. In line with its reduced ability to form amyloids (see Fig 3 and S4 Fig), the mechanical conformational polymorphism of the F5Y mutant is diminished, increasing the proportion of NM conformers (n = 109). (B) Immunodot blot showed that like toxic oligomeric intermediates of other amyloidogenic proteins, both full-length Orb2A, and to a lesser extent Orb2B, as well as their isolated PLDs, are recognized by the A11 antibody [45]. (C) A representative electron micrograph of aged Orb2A PLD shows the formation oligomers (asterisks) and typical unbranched amyloid fibers (arrows) resembling those of the full-length Orb2A (see Fig 1G) and pathological amyloids. Scale bar: 0.2 μm. Like pathological amyloids, those species are recognized by the fiber-specific OC monoclonal antibody [46]. For panels B and C, Aβ42 oligomers and fibers were used as positive controls for the A11 and OC antibodies, respectively, while ubiquitin was used as a negative control. The underlying data for panels in this figure can be found in S1 Data.