Robust Heteronuclear Correlations for Sub-milligram Protein in Ultrafast Magic-Angle Spinning Solid-State NMR.

Area of Science:

• Structural Biology
• Biophysics
• Analytical Chemistry

Background:

• Proton-detected solid-state NMR (ssNMR) is crucial for protein structure elucidation with limited sample amounts.
• Ultrafast magic-angle spinning (MAS) enhances ssNMR but diminishes traditional ¹³C-¹⁵N cross-polarization (CP) efficiency.
• Efficient heteronuclear correlations are vital for structural assignments in ssNMR.

Purpose of the Study:

• To develop a novel method to enhance ¹³C-¹⁵N correlation efficiency in ssNMR under ultrafast MAS.
• To overcome the limitations of traditional cross-polarization (CP) at high spinning speeds.
• To provide a more efficient tool for structural biology studies of proteins available in sub-milligram quantities.

Main Methods:

• Development and application of selective polarization between insensitive nuclei (SPINE) for heteronuclear ¹³C-¹⁵N transfer.
• Utilizing ultrafast magic-angle spinning (MAS) ssNMR spectroscopy.
• Validation of SPINE on diverse protein samples including GB1, MaMscL, SepF, and CcmL.

Main Results:

• SPINE significantly enhances ¹³C-¹⁵N correlation efficiency compared to CP.
• Observed gain factors of 1.75 for ¹³CA-¹⁵N and 1.9 for ¹³CO-¹⁵N transfers.
• Reduced experimental times by up to one-third (single CP) or one-tenth (dual CP).

Conclusions:

• SPINE is a robust and versatile method for improving ssNMR experiments.
• This technique facilitates faster and more efficient structural determination of proteins.
• SPINE is a valuable tool for advancing structural biology research, especially for challenging protein targets.