Magic Angle Spinning NMR Structure Determination of Proteins from Pseudocontact Shifts

Type
Article

Authors
Li, Jianping
Pilla, Kala Bharath
Li, Qingfeng
Zhang, Zhengfeng
Su, Xuncheng
Huber, Thomas
Yang, Jun

Online Publication Date
2013-05-24

Print Publication Date
2013-06-05

Date
2013-05-24

Abstract
Magic angle spinning solid-state NMR is a unique technique to study atomic-resolution structure of biomacromolecules which resist crystallization or are too large to study by solution NMR techniques. However, difficulties in obtaining sufficient number of long-range distance restraints using dipolar coupling based spectra hamper the process of structure determination of proteins in solid-state NMR. In this study it is shown that high-resolution structure of proteins in solid phase can be determined without the use of traditional dipolar-dipolar coupling based distance restraints by combining the measurements of pseudocontact shifts (PCSs) with Rosetta calculations. The PCSs were generated by chelating exogenous paramagnetic metal ions to a tag 4-mercaptomethyl-dipicolinic acid, which is covalently attached to different residue sites in a 56-residue immunoglobulin-binding domain of protein G (GB1). The long-range structural restraints with metal-nucleus distance of up to ∼20 Å are quantitatively extracted from experimentally observed PCSs, and these are in good agreement with the distances back-calculated using an X-ray structure model. Moreover, we demonstrate that using several paramagnetic ions with varied paramagnetic susceptibilities as well as the introduction of paramagnetic labels at different sites can dramatically increase the number of long-range restraints and cover different regions of the protein. The structure generated from solid-state NMR PCSs restraints combined with Rosetta calculations has 0.7 Å root-mean-square deviation relative to X-ray structure. © 2013 American Chemical Society.

Citation
Li J, Pilla KB, Li Q, Zhang Z, Su X, et al. (2013) Magic Angle Spinning NMR Structure Determination of Proteins from Pseudocontact Shifts. Journal of the American Chemical Society 135: 8294–8303. Available: http://dx.doi.org/10.1021/ja4021149.

Acknowledgements
J.Y. thanks Tatyana Polenova for valuable discussions. The authors thank Xu Zhang for help in solution NMR experiments. This work is supported by grants from the National Natural Science Foundation of China (21075133, 21173259, 21073101) and the National Basic Research Program of China (2009CB918600). We are thankful to King Abdulla University of Science and Technology (KAUST), Saudi Arabia, for providing access to the Blue Gene/P (Shaheen) supercomputer. T.H. acknowledges funding from the Australian Research Council, including a Future Fellowship (FT0991709) and project grant (DP120100561).

Publisher
American Chemical Society (ACS)

Journal
Journal of the American Chemical Society

DOI
10.1021/ja4021149

PubMed ID
23646876

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