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Determining methyl sidechain conformations in a CS-ROSETTA model using methyl 1H-13C residual dipolar couplings

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Modelling of protein structures based on backbone chemical shifts, using programs such as CS-ROSETTA, is becoming increasingly popular, especially for systems where few restraints are available or where homologous structures are already known. While the reliability of CS-ROSETTA calculations can be improved by incorporation of some additional backbone NMR data such as those afforded by residual dipolar couplings or minimal NOE data sets involving backbone amide protons, the sidechain conformations are largely modelled by statistical energy terms. Here, we present a simple method based on methyl residual dipolar couplings that can be used to determine the rotameric state of the threefold symmetry axis of methyl groups that occupy a single rotamer, determine rotameric distributions, and identify regions of high flexibility. The method is demonstrated for methyl side chains of a deletion variant of the human chaperone DNAJB6b.

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We thank Drs. James Baber, Jinfa Ying and Dan Garrett for technical support. This work was supported by the Intramural Program of the National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health (DK029023 to G.M.C.).

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Correspondence to Vitali Tugarinov or G. Marius Clore.

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Supplementary file 1 The pulse scheme used for recording the J-modulated interferograms (Fig. S1). Description of theoretical considerations involved in the interpretation of J-modulated interferograms of 13CH3 methyl groups in the presence of spin relaxation according to Eq. (2) (Fig. S2). A plot of values obtained for ΔST-DNAJB6b using the described analysis of J-modulated interferograms (Fig. S3). ‘Materials and Methods’ section describing the details of NMR sample preparation, NMR experiments and structure calculation protocols. (PDF 605 kb)

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Karamanos, T.K., Tugarinov, V. & Clore, G.M. Determining methyl sidechain conformations in a CS-ROSETTA model using methyl 1H-13C residual dipolar couplings. J Biomol NMR 74, 111–118 (2020).

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