Scholz Ingo, Jehle Stefan, Schmieder Peter, Hiller Matthias, Eisenmenger Frank, Oschkinat Hartmut, van Rossum Barth-Jan
Leibnizinstitut für Molekulare Pharmakologie, Berlin, Germany.
J Am Chem Soc. 2007 May 30;129(21):6682-3. doi: 10.1021/ja070849g. Epub 2007 May 9.
Scalar couplings between 13C spins can impair both resolution and sensitivity in 13C-labeled preparations. It is demonstrated that deconvolution of magic-angle-spinning NMR data with maximum entropy (MaxEnt) reconstruction allows the removal of splittings due to J-couplings without expenses in sensitivity. A combination of MaxEnt reconstruction in t2 with selective pulses in t1 produces fully J-resolved data in both dimensions. The possibility to obtain J-resolved 13C-13C data without compromising the sensitivity is particularly important for solid-state NMR of "difficult" biological samples, like membrane proteins, where sacrifices in signal-to-noise are fatal. The method is demonstrated using preparations of alpha-spectrin SH3 domain (62 residues) as small test system and of outermembrane protein G as example of a membrane protein with higher molecular weight (281 residues). Both preparations were obtained using [2-13C]-glycerol as the carbon source during the bacterial growth.
13C自旋之间的标量耦合会损害13C标记制剂的分辨率和灵敏度。结果表明，利用最大熵（MaxEnt）重建对魔角旋转NMR数据进行反卷积处理，可以去除由于J耦合引起的分裂，而不会损失灵敏度。在t2中进行MaxEnt重建与在t1中使用选择性脉冲相结合，可在两个维度上产生完全J分辨的数据。对于“难处理”的生物样品（如膜蛋白）的固态NMR而言，在不降低灵敏度的情况下获得J分辨的13C-13C数据的可能性尤为重要，因为在这些样品中牺牲信噪比是致命的。使用α-血影蛋白SH3结构域（62个残基）的制剂作为小型测试系统，并以外膜蛋白G作为较高分子量（281个残基）的膜蛋白示例来证明该方法。两种制剂均在细菌生长过程中使用[2-13C]-甘油作为碳源获得。
