Department of Chemistry and Ames Laboratory and ‡Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University , Ames, Iowa 50011, United States.
Biochemistry. 2014 May 6;53(17):2840-54. doi: 10.1021/bi500231b. Epub 2014 Apr 22.
The polysaccharide structure and dynamics in the primary cell wall of the model grass Brachypodium distachyon are investigated for the first time using solid-state nuclear magnetic resonance (NMR). While both grass and non-grass cell walls contain cellulose as the main structural scaffold, the former contains xylan with arabinose and glucuronic acid substitutions as the main hemicellulose, with a small amount of xyloglucan (XyG) and pectins, while the latter contains XyG as the main hemicellulose and significant amounts of pectins. We labeled the Brachypodium cell wall with (13)C to allow two-dimensional (2D) (13)C correlation NMR experiments under magic-angle spinning. Well-resolved 2D spectra are obtained in which the (13)C signals of cellulose, glucuronoarabinoxylan (GAX), and other matrix polysaccharides can be assigned. The assigned (13)C chemical shifts indicate that there are a large number of arabinose and xylose linkages in the wall, and GAX is significantly branched at the developmental stage of 2 weeks. 2D (13)C-(13)C correlation spectra measured with long spin diffusion mixing times indicate that the branched GAX approaches cellulose microfibrils on the nanometer scale, contrary to the conventional model in which only unbranched GAX can bind cellulose. The GAX chains are highly dynamic, with average order parameters of ~0.4. Biexponential (13)C T1 and (1)H T1ρ relaxation indicates that there are two dynamically distinct domains in GAX: the more rigid domain may be responsible for cross-linking cellulose microfibrils, while the more mobile domain may fill the interfibrillar space. This dynamic heterogeneity is more pronounced than that of the non-grass hemicellulose, XyG, suggesting that GAX adopts the mixed characteristics of XyG and pectins. Moderate differences in cellulose rigidity are observed between the Brachypodium and Arabidopsis cell walls, suggesting different effects of the matrix polysaccharides on cellulose. These data provide the first molecular-level structural information about the three-dimensional organization of the polysaccharides in the grass primary wall.
首次使用固态核磁共振（NMR）研究了模式草柳枝稷初生细胞壁中的多糖结构和动力学。虽然草类和非草类细胞壁都含有纤维素作为主要结构支架，但前者含有木聚糖，阿拉伯糖和葡萄糖醛酸取代作为主要半纤维素，少量木葡聚糖（XyG）和果胶，而后者含有 XyG 作为主要半纤维素和大量的果胶。我们用（13）C 标记柳枝稷细胞壁，以允许在魔角旋转下进行二维（2D）（13）C 相关 NMR 实验。得到了很好分辨的 2D 谱，其中可以分配纤维素、半乳糖阿拉伯木聚糖（GAX）和其他基质多糖的（13）C 信号。分配的（13）C 化学位移表明细胞壁中有大量的阿拉伯糖和木糖键，并且在 2 周的发育阶段，GAX 明显分支。用长自旋扩散混合时间测量的 2D（13）C-（13）C 相关谱表明，分支的 GAX 在纳米尺度上接近纤维素微纤维，与只有无支链 GAX 才能与纤维素结合的传统模型相反。GAX 链具有高度的动态性，平均顺序参数约为 0.4。双指数（13）C T1 和（1）H T1ρ弛豫表明 GAX 中有两个动态不同的域：更刚性的域可能负责交联纤维素微纤维，而更具流动性的域可能填充纤维间空间。这种动态异质性比非草类半纤维素 XyG 更为明显，表明 GAX 采用了 XyG 和果胶的混合特性。在柳枝稷和拟南芥细胞壁之间观察到纤维素刚性的适度差异，这表明基质多糖对纤维素有不同的影响。这些数据提供了关于草类初生壁中多糖三维组织的第一个分子水平结构信息。
