Selective detection of 13CH signals from the mixture of 13CH/13CH2/13CH3 groups in biomolecules with enhanced sensitivity

<title>Abstract</title> <p> The NMR spectra of ¹³ C-labeled biomolecules typically contain signals arising from ¹³ CH, ¹³ CH ₂ , and ¹³ CH ₃ groups. In many applications, ¹³ CH group is of the primary interest since it exhibits relatively simple spin system and relaxation behavior. However, the selective detection of ¹³ CH signals in the presence of ¹³ CH ₂ / ¹³ CH ₃ groups remains challenging, and many existing schemes are not well suited for biomolecular studies. In this work, we have developed an optimized pulse scheme, termed CH-selective ¹ H– ¹³ C HSQC, for selective detection of ¹³ CH signals from the mixtures containing ¹³ CH/ ¹³ CH ₂ / ¹³ CH ₃ groups. This method exploits one-bond ¹ <italic>J</italic> _CH scalar coupling evolution to efficiently suppress signals from ¹³ CH ₂ / ¹³ CH ₃ groups, while enhancing signal intensities of ¹³ CH resonances by exploiting cross-polarization (CP) based coherence transfer and preservation of equivalent pathways (PEP) methodology. This experiment offers distinct advantages over many other existing schemes developed for similar purposes, and is demonstrated using several protein samples. In addition, the experimental conditions to achieve optimal suppression of ¹³ CH ₂ / ¹³ CH ₃ moieties are systematically investigated and discussed. </p>