Fast magic angle spinning NMR with heteronucleus detection for resonance assignments and structural characterization of fully protonated proteins.
Heteronucleus-detected dipolar based correlation spectroscopy is established for assignments of ¹H, ¹³C, and ¹⁵N resonances and structural analysis in fully protonated proteins. We demonstrate that ¹³C detected 3D experiments are highly efficient and permit assignments of the majority of backbone resonances, as shown in an 89-residue dynein light chain 8, LC8 protein. With these experiments, we have resolved many ambiguities that were persistent in our previous studies using moderate MAS frequencies and lacking the ¹H dimension. The availability of ¹H isotropic chemical shifts measured with the heteronucleus-detected fast-MAS experiments presented here is essential for the accurate determination of the ¹H CSA tensors, which provide very useful structural probe. Finally, our results indicate that ¹³C detection in fast-MAS HETCOR experiments may be advantageous compared with ¹H detection as it yields datasets of significantly higher resolution in the ¹³C dimension than the ¹H detected HETCOR versions.
Department of Chemistry and Biochemistry, University of Delaware, Newark, DE, 19716, USA, cmguo@udel.edu.
Changmiao
C
Guo
Guangjin
G
Hou
Xingyu
X
Lu
Bernie
B
O'Hare
Jochem
J
Struppe
Tatyana
T
Polenova
D000595:Amino Acid Sequence; D056445:Cytoplasmic Dyneins / Q000737:chemistry; D029721:Drosophila Proteins / Q000737:chemistry; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011506:Proteins / Q000737:chemistry*; D011522:Protons
D029721:Drosophila Proteins; D011506:Proteins; D011522:Protons; D056445:Cytoplasmic Dyneins