Carbonyl carbon transverse relaxation dispersion measurements and ms-s timescale motion in a protein hydrogen bond network
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- 作者：Rieko Ishima ; James Baber ; John M. Louis and Dennis A. Torchia
- 关键词：chemical exchange ; conformational change ; CPMG ; NMR ; R2
- 刊名：Journal of Biomolecular NMR
- 出版时间：June, 2004
- 出版年：2004
- 期刊代码：76_09252738
- 类别：bio
- 卷：29
- 期：2
- 页码：187-198
- 数据来源：sp
摘要
A constant-time, Carr–Purcell–Meiboom–Gill (CPMG) transverse relaxation, R2, dispersion experiment for carbonyl carbons was designed and executed to detect s-ms time-scale dynamics of protein backbone carbonyl sites. Because of the large (ca. 55 Hz) C-C J-coupling, the carbonyl signal intensity is strongly modulated as the spacing between CPMG pulses is varied, in uniformly 13C enriched proteins, unless care is taken to minimize the perturbation of the C magnetization by the CPMG pulses. CPMG pulse trains consisting of either a band-selective pulse, such as RE-BURP, or rectangular (with an excitation null in the C region of the spectrum) pulses were employed in order to minimize C signal modulation by C-C J-coupling. The performance of these types of CPMG refocusing pulses was assessed by computer simulation, and by comparing dispersion profiles measured for (1) uniformly [13C,15N, 2H] (2H at non-labile hydrogen sites) labeled, and (2) uniformly 15N/selectively-13C labeled samples of HIV-1 protease bound to a potent inhibitor, DMP323. In addition, because the uniformly 13C/15N/2H labeled sample was well suited to measure 15N and 1H R2 dispersion as well as 13C dispersion, conformational exchange in the inter subunit -sheet hydrogen-bond network of the inhibitor-bound protease was elucidated using relaxation dispersion data of all three types of nuclei.