Under ultrafast magic-angle spinning (MAS), proton-detected solid-state nuclear magnetic resonance (ssNMR) has emerged as a powerful technique for elucidating structures from sub-milligram protein, where establishing 13C-15N correlations is essential. However, traditional 13C-15N cross-polarization (CP), which performs well at lower MAS frequencies, suffers diminished efficiency under ultrafast MAS conditions. To address this challenge, we have developed a robust method for selective polarization between insensitive nuclei (SPINE). This approach significantly enhances the efficiency of heteronuclear 13C-15N correlation compared to CP, achieving gain factors of 1.75 for 13CA-15N and 1.9 and 13CO-15N transfers. These enhancements can reduce the duration of current multi-dimensional experiments to approximately one-third of that required by 13C-15N CP and to approximately one-tenth when involving two 13C-15N transfers. The effectiveness of SPINE has been validated through experiments on four distinct proteins: the microcrystalline β1 immunoglobulin binding domain of protein G (GB1), the large-conductance mechanosensitive ion channel from Methanosarcina acetivorans (MaMscL), fibrillar septum-forming protein (SepF), and the vertex protein of the β-carboxysome shell (CcmL). These findings highlight the practical utility and versatility of SPINE in ssNMR spectroscopy, making it a valuable approach for structural biology applications.

中文翻译：

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超快魔角旋转固体 NMR 中蛋白质的稳健异核相关性


在超快魔角旋转 （MAS） 下，质子检测的固态核磁共振 （ssNMR） 已成为一种从亚毫克蛋白质中阐明结构的强大技术，其中建立 13C-15N 相关性至关重要。然而，传统的 13C-15N 交叉极化 （CP） 在较低的 MAS 频率下表现良好，但在超快 MAS 条件下效率降低。为了应对这一挑战，我们开发了一种强大的非敏感核之间选择性极化 （SPINE） 方法。与 CP 相比，这种方法显著提高了异核 13C-15N 相关性的效率，使 13CA-15N 和 1.9 和 13CO-15N 转移的增益因子达到 1.75。这些增强功能可以将当前多维实验的持续时间缩短到大约 13C-15N CP 所需时间的三分之一，在涉及两次 13C-15N 转移时减少到大约十分之一。SPINE 的有效性已通过对四种不同蛋白质的实验得到验证：蛋白 G 的微晶 β1 免疫球蛋白结合域 （GB1）、来自乙酰甲烷肉的大电导机械敏感离子通道 （MaMscL）、纤维隔膜形成蛋白 （SepF） 和β-羧基体壳的顶点蛋白 （CcmL）。这些发现突出了 SPINE 在 ssNMR 波谱学中的实际实用性和多功能性，使其成为结构生物学应用的宝贵方法。