Guo Chenyun, Tugarinov Vitali
Department of Chemistry and Biochemistry, University of Maryland, College Park, 20742, USA.
J Biomol NMR. 2009 Jan;43(1):21-30. doi: 10.1007/s10858-008-9285-8. Epub 2008 Nov 11.
A pair of HN-methyl NOESY experiments that are based on simultaneous TROSY-type detection of amide and methyl groups is described. The preservation of cross-peak symmetry in the simultaneous (1)H-(15)N/(13)CH(3) NOE spectra enables straightforward assignments of HN-methyl NOE cross-peaks in large and complex protein structures. The pulse schemes are designed to preserve the slowly decaying components of both (1)H-(15)N and methyl (13)CH(3) spin-systems in the course of indirect evolution (t (2)) and acquisition period (t (3)) of 3D NOESY experiments. The methodology has been tested on {U-[(15)N,(2)H]; Iledelta1-[(13)CH(3)]; Leu,Val-[(13)CH(3),(12)CD(3)]}-labeled 82-kDa enzyme Malate Synthase G (MSG). A straightforward procedure that utilizes the symmetry of NOE cross-peaks in the time-shared 3D NOE data sets allows unambiguous assignments of more than 300 HN-methyl interactions in MSG from a single 3D data set providing important structural restraints for derivation of the backbone global fold.
描述了一对基于酰胺和甲基同时进行TROSY型检测的HN-甲基NOESY实验。在同时进行的（1）H-（15）N/（13）CH（3）NOE谱中交叉峰对称性的保留，使得能够直接对大型复杂蛋白质结构中的HN-甲基NOE交叉峰进行归属。脉冲序列的设计目的是在3D NOESY实验的间接演化（t（2））和采集期（t（3））过程中保留（1）H-（15）N和甲基（13）CH（3）自旋系统的缓慢衰减成分。该方法已在{U-[（15）N，（2）H]；Ileδ1-[（13）CH（3）]；Leu，Val-[（13）CH（3），（12）CD（3）]}标记的82 kDa苹果酸合酶G（MSG）上进行了测试。利用分时3D NOE数据集中NOE交叉峰的对称性的直接程序，允许从单个3D数据集中明确归属MSG中超过300个HN-甲基相互作用，为推导主链全局折叠提供重要的结构限制。
