25-30 August 2019
Henry Ford Building
Europe/Berlin timezone

Including protons in solid-state NMR resonance assignment and secondary structure analysis for RNA polymerase II subunits Rpo4/7

Not scheduled
4h
Harnack House and Henry Ford Building

Harnack House and Henry Ford Building

Board: 474
Poster Posters

Speaker

Anahit Torosyan

Description

1H-detected solid-state NMR experiments with fast magic-angle-spinning (MAS) frequencies deliver 1H chemical shifts of proteins in solids, which enables their interpretation in terms of secondary structure. We present 1H and 13C-detected NMR spectra of the RNA polymerase1 subunit Rpo7 in complex with unlabeled Rpo4 and use the 13C, 15N and 1H chemical-shift values deduced from them to study the secondary structure of the protein in comparison to a known2 crystal structure. We applied the automated resonance assignment approach FLYA, including 1H solid-state chemical shifts, and show its performance in comparison to manual spectral assignment. Further assessing the secondary structure in comparison to the known crystal structure, our results confirm that 13C secondary chemical shifts (SCS) are a bona fide predictor of secondary structure elements. In cases where 13C chemical shifts are not available, secondary structure elements can be identified using the sum of 1Hα and 1HNN SCS, while using instead either only 1Hα or 1HN SCS or TALOS showed an increased uncertainty in the boundaries of observed secondary structure elements compared to the crystal structure2. We discuss the potential as well as pitfalls of secondary structure determination based on solid-state NMR including 1H chemical shifts.

[1] F. Werner & D. Grohmann. Nat. Rev. Microbiol. (2011) 9, 85–98.
[2] F. Todone, P. Brick, F. Werner, R.O. Weinzierl, S. Onesti, Mol Cell. (2001) 8(5), 1137-1143

Primary authors

Anahit Torosyan Thomas Wiegand Maarten Schledorn (Physical Chemistry, ETH Zurich, 8093 Zurich, Switzerland) Dr Daniel Klose (ETH Zurich) Prof. Peter Güntert (ETH Zürich) Mrs Anja Böckmann (Molecular Microbiology and Structural Biochemistry - UMR5086) Mr Beat H. Meier (Physical Chemistry, ETH Zürich)

Presentation Materials

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