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Self-consistent residual dipolar coupling based model-free analysis for the robust determination of nanosecond to microsecond protein dynamics.

Lakomek NA, Walter KF, Farès C, Lange OF, de Groot BL, Grubmüller H, Brüschweiler R, Munk A, Becker S, Meiler J, Griesinger C - J. Biomol. NMR (2008)

Bottom Line: For ubiquitin, the SCRM analysis yields an average RDC-derived order parameter of the NH vectors <S2(rdc)>0.72 +/- 0.02 compared to <S2(LS)> = 0.778 +/- 0.003 for the Lipari-Szabo order parameters, indicating that the inclusion of the supra-tau(c) window increases the averaged amplitude of mobility observed in the sub-supra-tau(c) window by about 34%.The backbone of Lys48, whose side chain is known to be involved in the poly-ubiquitylation process that leads to protein degradation, is very mobile on the supra-tau(c) time scale (S2(rdc)(NH) = 0.59 +/- 0.03), while it is inconspicuous (S2(LS)(NH)= 0.82) on the sub-tau(c) as well as on micros-ms relaxation dispersion time scales.The results of this work differ from previous RDC dynamics studies of ubiquitin in the sense that the results are essentially independent of structural noise providing a much more robust assessment of dynamic effects that underlie the RDC data.

View Article: PubMed Central - PubMed

Affiliation: Department for NMR-based Structural Biology, Max-Planck Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen, Germany.

ABSTRACT
Residual dipolar couplings (RDCs) provide information about the dynamic average orientation of inter-nuclear vectors and amplitudes of motion up to milliseconds. They complement relaxation methods, especially on a time-scale window that we have called supra-tau(c) (tau(c) < supra-tau(c) < 50 micros). Here we present a robust approach called Self-Consistent RDC-based Model-free analysis (SCRM) that delivers RDC-based order parameters-independent of the details of the structure used for alignment tensor calculation-as well as the dynamic average orientation of the inter-nuclear vectors in the protein structure in a self-consistent manner. For ubiquitin, the SCRM analysis yields an average RDC-derived order parameter of the NH vectors 0.72 +/- 0.02 compared to = 0.778 +/- 0.003 for the Lipari-Szabo order parameters, indicating that the inclusion of the supra-tau(c) window increases the averaged amplitude of mobility observed in the sub-supra-tau(c) window by about 34%. For the beta-strand spanned by residues Lys48 to Leu50, an alternating pattern of backbone NH RDC order parameter S2(rdc)(NH) = (0.59, 0.72, 0.59) was extracted. The backbone of Lys48, whose side chain is known to be involved in the poly-ubiquitylation process that leads to protein degradation, is very mobile on the supra-tau(c) time scale (S2(rdc)(NH) = 0.59 +/- 0.03), while it is inconspicuous (S2(LS)(NH)= 0.82) on the sub-tau(c) as well as on micros-ms relaxation dispersion time scales. The results of this work differ from previous RDC dynamics studies of ubiquitin in the sense that the results are essentially independent of structural noise providing a much more robust assessment of dynamic effects that underlie the RDC data.

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Sketch of the SCRM cycles: RDC-based model-free results are calculated using NH RDC data as well as inter-nuclear NH vectors for tensor calculation. Starting from a first structural model (e.g. the X-ray structure 1ubi with protons added with standard geometry), the inter-nuclear NH vector orientations are adjusted in each SCRM cycle towards the true dynamic average orientation. Consequently, the fit of the alignment tensor to the RDC data is improved in each step and as result the fit of the model-free results to the RDCs as well
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Fig1: Sketch of the SCRM cycles: RDC-based model-free results are calculated using NH RDC data as well as inter-nuclear NH vectors for tensor calculation. Starting from a first structural model (e.g. the X-ray structure 1ubi with protons added with standard geometry), the inter-nuclear NH vector orientations are adjusted in each SCRM cycle towards the true dynamic average orientation. Consequently, the fit of the alignment tensor to the RDC data is improved in each step and as result the fit of the model-free results to the RDCs as well

Mentions: The goal of the Self-Consistent RDC-based Model-free (SCRM) approach was to remove a possible bias due to the protein structure used for the initial determination of the alignment tensors. To this aim, we proceeded as follows. The first step of the SCRM method was the application of the standard RDC-based model-free analysis as described in (Lakomek et al. 2006): First, alignment tensors were calculated from the measured RDCs using the PALES (Zweckstetter and Bax 2000) or DipoCoup software (Meiler et al. 2000) and from the X-ray structure 1ubi (Ramage et al. 1994) to which protons were added with MOLMOL (Koradi et al. 1996). Naturally, the dynamically averaged orientations calculated from the RDC-based model-free approach will exhibit deviations from the orientations in the X-ray structure. In an iterative fashion, the NH vector orientations used for the tensor calculation were replaced by the resulting after each model-free analysis cycle (Fig. 1).Fig. 1


Self-consistent residual dipolar coupling based model-free analysis for the robust determination of nanosecond to microsecond protein dynamics.

Lakomek NA, Walter KF, Farès C, Lange OF, de Groot BL, Grubmüller H, Brüschweiler R, Munk A, Becker S, Meiler J, Griesinger C - J. Biomol. NMR (2008)

Sketch of the SCRM cycles: RDC-based model-free results are calculated using NH RDC data as well as inter-nuclear NH vectors for tensor calculation. Starting from a first structural model (e.g. the X-ray structure 1ubi with protons added with standard geometry), the inter-nuclear NH vector orientations are adjusted in each SCRM cycle towards the true dynamic average orientation. Consequently, the fit of the alignment tensor to the RDC data is improved in each step and as result the fit of the model-free results to the RDCs as well
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2480484&req=5

Fig1: Sketch of the SCRM cycles: RDC-based model-free results are calculated using NH RDC data as well as inter-nuclear NH vectors for tensor calculation. Starting from a first structural model (e.g. the X-ray structure 1ubi with protons added with standard geometry), the inter-nuclear NH vector orientations are adjusted in each SCRM cycle towards the true dynamic average orientation. Consequently, the fit of the alignment tensor to the RDC data is improved in each step and as result the fit of the model-free results to the RDCs as well
Mentions: The goal of the Self-Consistent RDC-based Model-free (SCRM) approach was to remove a possible bias due to the protein structure used for the initial determination of the alignment tensors. To this aim, we proceeded as follows. The first step of the SCRM method was the application of the standard RDC-based model-free analysis as described in (Lakomek et al. 2006): First, alignment tensors were calculated from the measured RDCs using the PALES (Zweckstetter and Bax 2000) or DipoCoup software (Meiler et al. 2000) and from the X-ray structure 1ubi (Ramage et al. 1994) to which protons were added with MOLMOL (Koradi et al. 1996). Naturally, the dynamically averaged orientations calculated from the RDC-based model-free approach will exhibit deviations from the orientations in the X-ray structure. In an iterative fashion, the NH vector orientations used for the tensor calculation were replaced by the resulting after each model-free analysis cycle (Fig. 1).Fig. 1

Bottom Line: For ubiquitin, the SCRM analysis yields an average RDC-derived order parameter of the NH vectors <S2(rdc)>0.72 +/- 0.02 compared to <S2(LS)> = 0.778 +/- 0.003 for the Lipari-Szabo order parameters, indicating that the inclusion of the supra-tau(c) window increases the averaged amplitude of mobility observed in the sub-supra-tau(c) window by about 34%.The backbone of Lys48, whose side chain is known to be involved in the poly-ubiquitylation process that leads to protein degradation, is very mobile on the supra-tau(c) time scale (S2(rdc)(NH) = 0.59 +/- 0.03), while it is inconspicuous (S2(LS)(NH)= 0.82) on the sub-tau(c) as well as on micros-ms relaxation dispersion time scales.The results of this work differ from previous RDC dynamics studies of ubiquitin in the sense that the results are essentially independent of structural noise providing a much more robust assessment of dynamic effects that underlie the RDC data.

View Article: PubMed Central - PubMed

Affiliation: Department for NMR-based Structural Biology, Max-Planck Institute for Biophysical Chemistry, Am Fassberg 11, Goettingen, Germany.

ABSTRACT
Residual dipolar couplings (RDCs) provide information about the dynamic average orientation of inter-nuclear vectors and amplitudes of motion up to milliseconds. They complement relaxation methods, especially on a time-scale window that we have called supra-tau(c) (tau(c) < supra-tau(c) < 50 micros). Here we present a robust approach called Self-Consistent RDC-based Model-free analysis (SCRM) that delivers RDC-based order parameters-independent of the details of the structure used for alignment tensor calculation-as well as the dynamic average orientation of the inter-nuclear vectors in the protein structure in a self-consistent manner. For ubiquitin, the SCRM analysis yields an average RDC-derived order parameter of the NH vectors 0.72 +/- 0.02 compared to = 0.778 +/- 0.003 for the Lipari-Szabo order parameters, indicating that the inclusion of the supra-tau(c) window increases the averaged amplitude of mobility observed in the sub-supra-tau(c) window by about 34%. For the beta-strand spanned by residues Lys48 to Leu50, an alternating pattern of backbone NH RDC order parameter S2(rdc)(NH) = (0.59, 0.72, 0.59) was extracted. The backbone of Lys48, whose side chain is known to be involved in the poly-ubiquitylation process that leads to protein degradation, is very mobile on the supra-tau(c) time scale (S2(rdc)(NH) = 0.59 +/- 0.03), while it is inconspicuous (S2(LS)(NH)= 0.82) on the sub-tau(c) as well as on micros-ms relaxation dispersion time scales. The results of this work differ from previous RDC dynamics studies of ubiquitin in the sense that the results are essentially independent of structural noise providing a much more robust assessment of dynamic effects that underlie the RDC data.

Show MeSH