Asakura Tetsuo, Okonogi Michi, Nakazawa Yasumoto, Yamauchi Kazuo
Department of Biotechnology, Tokyo University of Agriculture and Technology, 2-24-16 Nakacho, Koganei, Tokyo 184-8588, Japan.
J Am Chem Soc. 2006 May 10;128(18):6231-8. doi: 10.1021/ja060251t.
The structural analysis of natural protein fibers with mixed parallel and antiparallel beta-sheet structures by solid-state NMR is reported. To obtain NMR parameters that can characterize these beta-sheet structures, (13)C solid-state NMR experiments were performed on two alanine tripeptide samples: one with 100% parallel beta-sheet structure and the other with 100% antiparallel beta-sheet structure. All (13)C resonances of the tripeptides could be assigned by a comparison of the methyl (13)C resonances of Ala(3) with different [3-(13)C]Ala labeling schemes and also by a series of RFDR (radio frequency driven recoupling) spectra observed by changing mixing times. Two (13)C resonances observed for each Ala residue could be assigned to two nonequivalent molecules per unit cell. Differences in the (13)C chemical shifts and (13)C spin-lattice relaxation times (T(1)) were observed between the two beta-sheet structures. Especially, about 3 times longer T(1) values were obtained for parallel beta-sheet structure as compared to those of antiparallel beta-sheet structure, which could be explicable by the difference in the hydrogen-bond networks of both structures. This very large difference in T(1) becomes a good measure to differentiate between parallel or antiparallel beta-sheet structures. These differences in the NMR parameters found for the tripeptides may be applied to assign the parallel and antiparallel beta-sheet (13)C resonances in the asymmetric and broad methyl spectra of [3-(13)C]Ala silk protein fiber of a wild silkworm, Samia cynthia ricini.
报道了通过固态核磁共振对具有混合平行和反平行β-折叠结构的天然蛋白质纤维进行的结构分析。为了获得能够表征这些β-折叠结构的核磁共振参数，对两个丙氨酸三肽样品进行了¹³C固态核磁共振实验：一个具有100%的平行β-折叠结构，另一个具有100%的反平行β-折叠结构。通过比较不同[³-(¹³)C]Ala标记方案下Ala(3)的甲基¹³C共振，以及通过改变混合时间观察到的一系列射频驱动重新耦合（RFDR）光谱，可以确定三肽的所有¹³C共振。每个Ala残基观察到的两个¹³C共振可归属于每个晶胞中的两个不等价分子。在两种β-折叠结构之间观察到¹³C化学位移和¹³C自旋晶格弛豫时间（T(1)）的差异。特别是，与反平行β-折叠结构相比，平行β-折叠结构的T(1)值大约长3倍，这可以通过两种结构氢键网络的差异来解释。T(1)的这种非常大的差异成为区分平行或反平行β-折叠结构的一个很好的指标。在三肽中发现的这些核磁共振参数差异，可应用于确定野生蚕蓖麻蚕的[³-(¹³)C]Ala丝蛋白纤维不对称且宽泛的甲基光谱中的平行和反平行β-折叠¹³C共振。
