Combining DNP NMR with segmental and specific labeling to study a yeast prion protein strain that is not parallel in-register

Kendra K. Frederick, Vladimir K. Michaelis, Marc A. Caporini, Loren B. Andreas, Galia T. Debelouchina, Robert G. Griffin, Susan Lindquist

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

The yeast prion protein Sup35NM is a self-propagating amyloid. Despite intense study, there is no consensus on the organization of monomers within Sup35NM fibrils. Some studies point to a â-helical arrangement, whereas others suggest a parallel inregister organization. Intermolecular contacts are often determined by experiments that probe long-range heteronuclear contacts for fibrils templated from a 1:1 mixture of 13C- and 15N-labeled monomers. However, for Sup35NM, like many large proteins, chemical shift degeneracy limits the usefulness of this approach. Segmental and specific isotopic labeling reduce degeneracy, but experiments to measure long-range interactions are often too insensitive. To limit degeneracy and increase experimental sensitivity, we combined specific and segmental isotopic labeling schemes with dynamic nuclear polarization (DNP) NMR. Using this combination, we examined an amyloid form of Sup35NM that does not have a parallel in-register structure. The combination of a small number of specific labels with DNP NMR enables determination of architectural information about polymeric protein systems.

Original languageEnglish (US)
Pages (from-to)3642-3647
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume114
Issue number14
DOIs
StatePublished - Apr 4 2017

Keywords

  • Amyloid
  • Dynamic nuclear polarization
  • Solid-state NMR
  • Sup35
  • [PSI] prion

ASJC Scopus subject areas

  • General

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