Dynamics of Lysine Side-Chain Amino Groups in a Protein Studied by Heteronuclear 1H鈭?sup>15N NMR Spectroscopy

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• 作者：Alexandre Esadze ; Da-Wei Li ; Tianzhi Wang ; Rafael Br眉schweiler ; Junji Iwahara
• 刊名：Journal of the American Chemical Society
• 出版年：2011
• 出版时间：February 2, 2011
• 年：2011
• 卷：133
• 期：4
• 页码：909-919
• 全文大小：1034K
• 年卷期：v.133,no.4(February 2, 2011)
• ISSN：1520-5126

文摘

Despite their importance in macromolecular interactions and functions, the dynamics of lysine side-chain amino groups in proteins are not well understood. In this study, we have developed the methodology for the investigations of the dynamics of lysine NH₃⁺ groups by NMR spectroscopy and computation. By using ¹H鈭?sup>15N heteronuclear correlation experiments optimized for ¹⁵NH₃⁺ moieties, we have analyzed the dynamic behavior of individual lysine NH₃⁺ groups in human ubiquitin at 2 掳C and pH 5. We modified the theoretical framework developed previously for CH₃ groups and used it to analyze ¹⁵N relaxation data for the NH₃⁺ groups. For six lysine NH₃⁺ groups out of seven in ubiquitin, we have determined model-free order parameters, correlation times for bond rotation, and reorientation of the symmetry axis occurring on a pico- to nanosecond time scale. From CPMG relaxation dispersion experiment for lysine NH₃⁺ groups, slower dynamics occurring on a millisecond time scale have also been detected for Lys27. The NH₃⁺ groups of Lys48, which plays a key role as the linkage site in ubiquitination for proteasomal degradation, was found to be highly mobile with the lowest order parameter among the six NH₃⁺ groups analyzed by NMR. We compared the experimental order parameters for the lysine NH₃⁺ groups with those from a 1 渭s molecular dynamics simulation in explicit solvent and found good agreement between the two. Furthermore, both the computer simulation and the experimental correlation times for the bond rotations of NH₃⁺ groups suggest that their hydrogen bonding is highly dynamic with a subnanosecond lifetime. This study demonstrates the utility of combining NMR experiment and simulation for an in-depth characterization of the dynamics of these functionally most important side-chains of ubiquitin.