CACA-TOCSY with alternate 13C-12C labeling: a 13Calpha direct detection experiment for mainchain resonance assignment, dihedral angle information, and amino acid type identification

J Biomol NMR. 2010 May;47(1):55-63. doi: 10.1007/s10858-010-9410-3. Epub 2010 Apr 10.

Abstract

We present a (13)C direct detection CACA-TOCSY experiment for samples with alternate (13)C-(12)C labeling. It provides inter-residue correlations between (13)C(alpha) resonances of residue i and adjacent C(alpha)s at positions i - 1 and i + 1. Furthermore, longer mixing times yield correlations to C(alpha) nuclei separated by more than one residue. The experiment also provides C(alpha)-to-sidechain correlations, some amino acid type identifications and estimates for psi dihedral angles. The power of the experiment derives from the alternate (13)C-(12)C labeling with [1,3-(13)C] glycerol or [2-(13)C] glycerol, which allows utilizing the small scalar (3)J(CC) couplings that are masked by strong (1)J(CC) couplings in uniformly (13)C labeled samples.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acids / chemistry*
  • Carbon Isotopes / chemistry*
  • Carbon Isotopes / metabolism
  • Deuterium / chemistry
  • Deuterium / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Glycerol / metabolism
  • Nerve Tissue Proteins / chemistry*
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism
  • Nuclear Magnetic Resonance, Biomolecular / methods*
  • Protein Conformation
  • Protein Subunits
  • Recombinant Fusion Proteins / chemistry*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism

Substances

  • Amino Acids
  • Carbon Isotopes
  • G-substrate
  • Nerve Tissue Proteins
  • Protein Subunits
  • Recombinant Fusion Proteins
  • Deuterium
  • Glycerol