Carbon-Detected NMR Experiments To Investigate Structure and Dynamics of Biological Macromolecules

Abstract

We have started to develop new NMR pulse sequences that detect carbon magnetization during the acquisition period. These experiments have become possible with the recent introduction of cryogenic probe heads. We show that a careful design of these carbon-detected experiments can at least partially compensate for the inherent lower sensitivity of carbon detection compared to proton detection. We discuss potential applications of carbon detection and demonstrate a deconvolution technique that removes the effects of carbon-carbon couplings from the spectra.

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

“…The carbon-detected version of the HCC-TOCSY-experiment (Serber et al 2001) developed for the assignment of protein side chains was one of the first applications of the new generation of 13 C-direct detected NMR experiments. Variants of this method were applied to large deuterated proteins (Eletsky et al 2003) and to selectively protonated proteins for the assignment of the methyl groups (Hu et al 2006; Jordan et al 2006).…”

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

“…The carbon-detected version of the HCC-TOCSY-experiment (Serber et al 2001) developed for the assignment of protein side chains was one of the first applications of the new generation of 13 C-direct detected NMR experiments. Variants of this method were applied to large deuterated proteins (Eletsky et al 2003) and to selectively protonated proteins for the assignment of the methyl groups (Hu et al 2006; Jordan et al 2006).…”

13 C Direct Detected NMR for Challenging Systems

“…These experiments were thus generally referred to as "exclusively heteronuclear" to indicate that only heteronuclei were frequency labeled in all dimensions, regardless of the starting polarization source exploited. 24,28,53,59 Considering for the moment backbone nuclear spins, the proton polarization source can be provided by amide protons ( 1 H N ) as well as by aliphatic protons ( 1 H α ) with advantages and disadvantages that depend on the properties of these two nuclei (Figure 2). Amide protons, which can be easily correlated to the directly bound nitrogen and then to carbonyl carbon nuclei through the 1 J HN and 1 J C′N , respectively, are influenced by solvent exchange that can become so pronounced that coherence transfer becomes inefficient.…”

Driving force of biomolecular liquid–liquid phase separation probed by nuclear magnetic resonance spectroscopy

“…The challenge is that LCDs are intrinsically flexible; thus, they can interconvert between various conformational states (Jensen et al 2014 ). Another challenge is the signal degeneration of amide protons due to the low sequence complexity and disordered structures; in such cases, 13 C or 15 N direct detection experiments can be performed (Bermel et al 2006a ; Eletsky et al 2003 ; Pervushin and Eletsky 2003 ; Serber et al 2000 , 2001 ; Takeuchi et al 2008 ). The spatial arrangement of the 13 C or 15 N coil was optimized to enhance the sensitivity of these two nuclei (Kovacs et al 2005 ).…”

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

“…The carbon-detected version of the HCC-TOCSY-experiment (Serber et al 2001) developed for the assignment of protein side chains was one of the first applications of the new generation of 13 C-direct detected NMR experiments. Variants of this method were applied to large deuterated proteins (Eletsky et al 2003) and to selectively protonated proteins for the assignment of the methyl groups (Hu et al 2006; Jordan et al 2006).…”

13 C Direct Detected NMR for Challenging Systems

“…These experiments were thus generally referred to as "exclusively heteronuclear" to indicate that only heteronuclei were frequency labeled in all dimensions, regardless of the starting polarization source exploited. 24,28,53,59 Considering for the moment backbone nuclear spins, the proton polarization source can be provided by amide protons ( 1 H N ) as well as by aliphatic protons ( 1 H α ) with advantages and disadvantages that depend on the properties of these two nuclei (Figure 2). Amide protons, which can be easily correlated to the directly bound nitrogen and then to carbonyl carbon nuclei through the 1 J HN and 1 J C′N , respectively, are influenced by solvent exchange that can become so pronounced that coherence transfer becomes inefficient.…”

Driving force of biomolecular liquid–liquid phase separation probed by nuclear magnetic resonance spectroscopy

“…The challenge is that LCDs are intrinsically flexible; thus, they can interconvert between various conformational states (Jensen et al 2014 ). Another challenge is the signal degeneration of amide protons due to the low sequence complexity and disordered structures; in such cases, 13 C or 15 N direct detection experiments can be performed (Bermel et al 2006a ; Eletsky et al 2003 ; Pervushin and Eletsky 2003 ; Serber et al 2000 , 2001 ; Takeuchi et al 2008 ). The spatial arrangement of the 13 C or 15 N coil was optimized to enhance the sensitivity of these two nuclei (Kovacs et al 2005 ).…”

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

“…The carbon-detected version of the HCC-TOCSY-experiment (Serber et al 2001) developed for the assignment of protein side chains was one of the first applications of the new generation of 13 C-direct detected NMR experiments. Variants of this method were applied to large deuterated proteins (Eletsky et al 2003) and to selectively protonated proteins for the assignment of the methyl groups (Hu et al 2006; Jordan et al 2006).…”

13 C Direct Detected NMR for Challenging Systems

“…These experiments were thus generally referred to as "exclusively heteronuclear" to indicate that only heteronuclei were frequency labeled in all dimensions, regardless of the starting polarization source exploited. 24,28,53,59 Considering for the moment backbone nuclear spins, the proton polarization source can be provided by amide protons ( 1 H N ) as well as by aliphatic protons ( 1 H α ) with advantages and disadvantages that depend on the properties of these two nuclei (Figure 2). Amide protons, which can be easily correlated to the directly bound nitrogen and then to carbonyl carbon nuclei through the 1 J HN and 1 J C′N , respectively, are influenced by solvent exchange that can become so pronounced that coherence transfer becomes inefficient.…”

Driving force of biomolecular liquid–liquid phase separation probed by nuclear magnetic resonance spectroscopy

“…The challenge is that LCDs are intrinsically flexible; thus, they can interconvert between various conformational states (Jensen et al 2014 ). Another challenge is the signal degeneration of amide protons due to the low sequence complexity and disordered structures; in such cases, 13 C or 15 N direct detection experiments can be performed (Bermel et al 2006a ; Eletsky et al 2003 ; Pervushin and Eletsky 2003 ; Serber et al 2000 , 2001 ; Takeuchi et al 2008 ). The spatial arrangement of the 13 C or 15 N coil was optimized to enhance the sensitivity of these two nuclei (Kovacs et al 2005 ).…”