Sample optimization and identification of signal patterns of amino acid side chains in 2D RFDR spectra of the α-spectrin SH3 domain
Sample optimization and identification of signal patterns of amino acid side chains in 2D RFDR spectra of the α-spectrin SH3 domain
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DOI：
10.1006/jmre.2000.2029
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发表时间：
2000-04-01
影响因子：
2.2
通讯作者：
Oschkinat, H
中科院分区：
文献类型：
--
作者：
Pauli, J;van Rossum, B;Oschkinat, H
未来用固态CPP-MAS核磁共振对蛋白质的结构研究将依赖于均匀标记的蛋白质样品显示良好的分辨率的光谱。用四种不同的方法产生固体α-光谱蛋白SH3结构域的核磁共振样品，并比较了它们的C-13 CPMAS谱。由沉淀产生的[u-C-13，N-15]标记样品的光谱显示很窄的C-13信号和可分辨的标量碳-碳耦合，选择性标记的[70%3-C-13]丙氨酸富集型SH3样品的三个丙氨酸CP信号的线宽为16-19赫兹。在[u-C-13，N-15]标记的SH3样品的2D C-13-C-13RFDR谱中，确定了异亮氨酸、所有脯氨酸、缬氨酸、丙氨酸和丝氨酸以及四种苏氨酸中的三种的信号模式。将发现的信号模式的C-13化学位移与在溶液中获得的C-13赋值进行比较，发现了一个有趣的匹配。(C)2000年学术出版社。
Future structural investigations of proteins by solid-state CPP-MAS NMR will rely on uniformly labeled protein samples showing spectra with an excellent resolution. NMR samples of the solid alpha-spectrin SH3 domain were generated in four different ways, and their C-13 CPMAS spectra were compared. The spectrum of a [u-C-13, N-15]-labeled sample generated by precipitation shows very narrow C-13 signals and resolved scalar carbon-carbon couplings, Linewidths of 16-19 Hz were found for the three alanine CP signals of a selectively labeled [70% 3-C-13]alanine-enriched SH3 sample. The signal pattern of the isoleucine, of all prolines, valines, alanines, and serines, and of three of the four threonines were identified in 2D C-13-C-13 RFDR spectra of the [u-C-13, N-15]-labeled SH3 sample. A comparison of the C-13 chemical shifts of the found signal patterns with the C-13 assignment obtained in solution shows an intriguing match. (C) 2000 Academic Press.