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A straightforward method for stereospecific assignment of val and leu prochiral methyl groups by solid-state NMR: Scrambling in the [2-13C]Glucose labeling scheme.

MPS-Authors
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Lv,  G.
Department of NMR-Based Structural Biology, MPI for biophysical chemistry, Max Planck Society;

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Faßhuber,  H. K.
Research Group of Solid-State NMR, MPI for Biophysical Chemistry, Max Planck Society;

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Loquet,  A.
Research Group of Solid-State NMR, MPI for Biophysical Chemistry, Max Planck Society;

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Demers,  J. P.
Research Group of Solid-State NMR, MPI for Biophysical Chemistry, Max Planck Society;

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Vijayan,  V.
Research Group of Solid-State NMR, MPI for Biophysical Chemistry, Max Planck Society;

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Giller,  K.
Department of NMR-Based Structural Biology, MPI for biophysical chemistry, Max Planck Society;

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Becker,  S.
Department of NMR-Based Structural Biology, MPI for biophysical chemistry, Max Planck Society;

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Lange,  A.
Research Group of Solid-State NMR, MPI for Biophysical Chemistry, Max Planck Society;

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Fulltext (public)

1694970.pdf
(Publisher version), 984KB

Supplementary Material (public)

1694970-Suppl.doc
(Supplementary material), 3MB

Citation

Lv, G., Faßhuber, H. K., Loquet, A., Demers, J. P., Vijayan, V., Giller, K., et al. (2013). A straightforward method for stereospecific assignment of val and leu prochiral methyl groups by solid-state NMR: Scrambling in the [2-13C]Glucose labeling scheme. Journal of Magnetic Resonance, 228, 45-49. doi:10.1016/j.jmr.2012.12.017.


Cite as: http://hdl.handle.net/11858/00-001M-0000-000E-E51C-6
Abstract
The unambiguous stereospecific assignment of the prochiral methyl groups in Val and Leu plays an important role in the structural investigation of proteins by NMR. Here, we present a straightforward method for their stereospecific solid-state NMR assignment based on [2-13C]Glucose ([2-13C]Glc) as the sole carbon source during protein expression. The approach is fundamentally based on the stereo-selective biosynthetic pathway of Val and Leu, and the co-presence of [2-13C]pyruvate produced mainly by glycolysis and [3-13C]/[1,3-13C]pyruvate most probably formed through scrambling in the pentose phosphate pathway. As a consequence, the isotope spin pairs 13Cβ-13Cγ2 and 13Cα-13Cγ1 in Val, and 13Cγ-13Cδ2 and 13Cβ-13Cδ1 in Leu are obtained. The approach is successfully demonstrated with the stereospecific assignment of the methyl groups of Val and Leu of type 3 secretion system PrgI needles and microcrystalline ubiquitin.