Methods for sequential resonance assignment in solid, uniformly 13C, 15N labelled peptides: quantification and...

Area of Science:

• Solid-state Nuclear Magnetic Resonance (NMR) spectroscopy
• Structural biology
• Biophysical chemistry

Background:

• Resonance assignment in solid-state NMR is crucial for determining polypeptide structures.
• Traditional methods can be time-consuming and complex.
• Development of efficient polarization transfer techniques is essential.

Purpose of the Study:

• To demonstrate the application of adiabatic polarization-transfer experiments for resonance assignment in solid-state polypeptides.
• To compare the effectiveness of DREAM and TOBSY sequences for homonuclear correlation.
• To achieve complete sequence-specific assignment of 13C and 15N resonances in antamanide.

Main Methods:

• Utilized adiabatic polarization-transfer experiments, including DREAM and TOBSY sequences.
• Employed uniformly 13C-15N-labelled antamanide for solid-state NMR studies.
• Combined heteronuclear transfer (adiabatic-passage Hartmann-Hahn) with homonuclear transfer (DREAM, rotational-resonance tickling) in multi-dimensional experiments.

Main Results:

• The DREAM sequence achieved complete assignment of C(alpha) and aliphatic side-chain 13C resonances to amino acid types.
• The TOBSY experiment with P9(12)1 sequence provided identical homonuclear correlation information.
• A series of adiabatic triple-resonance experiments led to near-complete sequence-specific assignment of 13C and 15N resonances.

Conclusions:

• Adiabatic polarization-transfer experiments are highly effective for resonance assignment in solid-state polypeptides.
• Both DREAM and TOBSY sequences offer efficient homonuclear correlation for structural elucidation.
• These advanced NMR techniques significantly enhance the speed and completeness of structural determination for biomolecules.