HN-NCA heteronuclear TOCSY-NH experiment for (1)H(N) and (15)N sequential correlations in ((13)C, (15)N) labelled intrinsically disordered proteins.
A simple triple resonance NMR experiment that leads to the correlation of the backbone amide resonances of each amino acid residue 'i' with that of residues 'i-1' and 'i+1' in ((13)C, (15)N) labelled intrinsically disordered proteins (IDPs) is presented. The experimental scheme, {HN-NCA heteronuclear TOCSY-NH}, exploits the favourable relaxation properties of IDPs and the presence of (1) J CαN and (2) J CαN couplings to transfer the (15)N x magnetisation from amino acid residue 'i' to adjacent residues via the application of a band-selective (15)N-(13)C(α) heteronuclear cross-polarisation sequence of ~100 ms duration. Employing non-uniform sampling in the indirect dimensions, the efficacy of the approach has been demonstrated by the acquisition of 3D HNN chemical shift correlation spectra of α-synuclein. The experimental performance of the RF pulse sequence has been compared with that of the conventional INEPT-based HN(CA)NH pulse scheme. As the availability of data from both the HCCNH and HNN experiments will make it possible to use the information extracted from one experiment to simplify the analysis of the data of the other and lead to a robust approach for unambiguous backbone and side-chain resonance assignments, a time-saving strategy for the simultaneous collection of HCCNH and HNN data is also described.
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.|Institute of Biochemistry/Biotechnology, Martin Luther University Halle-Wittenberg, Kurt-Mothes-Strasse 3, 06120, Halle/Salle, Germany.
Christoph
C
Wiedemann
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.
Nishit
N
Goradia
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.
Sabine
S
Häfner
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.|Department of Physics, Faculty of Science, Ubon Ratchathani University, Ubon Ratchathani, 34190, Thailand.
Christian
C
Herbst
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.
Matthias
M
Görlach
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany.
Oliver
O
Ohlenschläger
Research Group Biomolecular NMR Spectroscopy, Leibniz Institute for Age Research, Fritz Lipmann Institute, Beutenbergstr. 11, 07745, Jena, Germany. raman@fli-leibniz.de.
Ramadurai
R
Ramachandran
D064267:Intrinsically Disordered Proteins / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*
D064267:Intrinsically Disordered Proteins
Heteronuclear cross polarisation; Intrinsically disordered protein region; NMR spectroscopy; Non-uniform sampling; Sequential resonance assignment