Structure Calculation from Unambiguous Long-Range Amide and Methyl H-1-H-1 Distance Restraints for a Microcrystalline Protein with MAS Solid-State NMR Spectroscopy

Rasmus Linser, Benjamin Bardiaux, Victoria Higman, Uwe Fink, Bernd Reif*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

106 Citations (Scopus)

Abstract

Magic-angle spinning (MAS) solid-state NMR becomes an increasingly important tool for the determination of structures of membrane proteins and amyloid fibrils. Extensive deuteration of the protein allows multidimensional experiments with exceptionally high sensitivity and resolution to be obtained. Here we present an experimental strategy to measure highly unambiguous spatial correlations for distances up to 13 angstrom. Two complementary three-dimensional experiments, or alternatively a four-dimensional experiment, yield highly unambiguous cross-peak assignments, which rely on four encoded chemical shift dimensions. Correlations to residual aliphatic protons are accessible via synchronous evolution of the N-15 and C-13 chemical shifts, which encode valuable amide-methyl distance restraints. On average, we obtain six restraints per residue. Importantly, 50% of all restraints correspond to long-range distances between residues i and j with vertical bar i-j vertical bar > 5, which are of particular importance in structure calculations. Using ARIA, we calculate a high-resolution structure for the microcrystalline 7.2 kDa alpha-spectrin SH3 domain with a backbone precision of similar to 1.1 angstrom.

Original languageEnglish
Pages (from-to)5905-5912
Number of pages8
JournalJournal of the American Chemical Society
Volume133
Issue number15
DOIs
Publication statusPublished - 20 Apr 2011

Keywords

  • RESOLUTION
  • ROTATING SOLIDS
  • PERDEUTERATED PROTEINS
  • AUTOMATED NOE ASSIGNMENT
  • ANGLE-SPINNING NMR
  • SH3 DOMAIN
  • 3D
  • C-13
  • BACKBONE
  • SENSITIVITY ENHANCEMENT

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