Utilization of paramagnetic relaxation enhancements for high-resolution NMR structure determination of a soluble loop-rich protein with sparse NOE distance restraints
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NMR structure determination of soluble proteins depends in large part on distance restraints derived from NOE. In this study, we examined the impact of paramagnetic relaxation enhancement (PRE)-derived distance restraints on protein structure determination. A high-resolution structure of the loop-rich soluble protein Sin1 could not be determined by conventional NOE-based procedures due to an insufficient number of NOE restraints. By using the 867 PRE-derived distance restraints obtained from the NOE-based structure determination procedure, a high-resolution structure of Sin1 could be successfully determined. The convergence and accuracy of the determined structure were improved by increasing the number of PRE-derived distance restraints. This study demonstrates that PRE-derived distance restraints are useful in the determination of a high-resolution structure of a soluble protein when the number of NOE constraints is insufficient.
KeywordsParamagnetic relaxation enhancement PRE Structure determination Soluble protein RDC Sin1
We thank Dr. Peter Güntert for letting us use CYANA 3.95. We thank Momoko Yoneyama, and Yuki Nishigaya for sample preparation and mass spectrometry measurements, respectively. This work was supported in part by Grants from MEXT/JSPS KAKENHI, TPRP, Platform for Drug Discovery, Informatics, and Structural Life Science to K.S., T.F. and C.K., and Grants-in-Aid for JSPS Fellows to K.F. and S.K.
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