Progress in FY2013 was in the following areas: LOW COMPLEXITY SEQUENCES. We have initiated a collaboration with the Prof. Steven McKnight of UT Southwestern to apply solid state NMR and electron microscopy to studies of homo- and hetero-association of proteins that contain "low complexity sequence" (LCS) domains. A wide variety of LCS-containing proteins, some involved in mRNA export and localization of gene expression, have been identified in Prof. McKnight's lab and implicated in formation of intracellular granules that appear to be comprised of amyloid-like fibrils. Solid state NMR measurements have been performed on fibrils formed by low complexity sequences of hnRNPA2 and FUS. 3D solid state NMR spectra of hnRNPA2 fibrils indicate polymorphism, as all NMR signals can not be assigned to a single copy of the hnRNPA2 sequence. Additional solid state NMR measurements indicate in-register parallel beta-sheets. 2D solid state NMR spectra of FUS fibrils are better resolved than those of hnRNPA2, suggesting a single predominant fibril structure. Remarkably, although the FUS low complexity sequence contains many Tyr residues, separated by 6- to 8-residue segments comprised mostly of Gly, Ser, Thr, and Gln residues, signals from Tyr residues are weak in solid state NMR spectra. In contrast, signals from Tyr residues are strong in NMR spectra recorded with solution NMR techniques. Thus, the immobilized core FUS fibrils appears to contain multiple short segments (probably in beta-sheets) that are separated by long, highly flexible loops. NMR chemical shift assignments are in progress, with the goal of developing a full structural model for FUS fibrils.

Project Start
Project End
Budget Start
Budget End
Support Year
5
Fiscal Year
2013
Total Cost
$373,103
Indirect Cost
City
State
Country
Zip Code
Tycko, Robert; Wickner, Reed B (2013) Molecular structures of amyloid and prion fibrils: consensus versus controversy. Acc Chem Res 46:1487-96
Shewmaker, Frank; Kryndushkin, Dmitry; Chen, Bo et al. (2009) Two prion variants of Sup35p have in-register parallel beta-sheet structures, independent of hydration. Biochemistry 48:5074-82
Chen, Bo; Thurber, Kent R; Shewmaker, Frank et al. (2009) Measurement of amyloid fibril mass-per-length by tilted-beam transmission electron microscopy. Proc Natl Acad Sci U S A 106:14339-44
Shewmaker, Frank; McGlinchey, Ryan P; Thurber, Kent R et al. (2009) The functional curli amyloid is not based on in-register parallel beta-sheet structure. J Biol Chem 284:25065-76
Shewmaker, Frank; Ross, Eric D; Tycko, Robert et al. (2008) Amyloids of shuffled prion domains that form prions have a parallel in-register beta-sheet structure. Biochemistry 47:4000-7
Wickner, Reed B; Dyda, Fred; Tycko, Robert (2008) Amyloid of Rnq1p, the basis of the [PIN+] prion, has a parallel in-register beta-sheet structure. Proc Natl Acad Sci U S A 105:2403-8