H-1-N-15 HETERONUCLEAR NMR-STUDIES OF ESCHERICHIA-COLI THIOREDOXIN IN SAMPLES ISOTOPICALLY LABELED BY RESIDUE TYPE
H-1-N-15 HETERONUCLEAR NMR-STUDIES OF ESCHERICHIA-COLI THIOREDOXIN IN SAMPLES ISOTOPICALLY LABELED BY RESIDUE TYPE
复制标题
DOI：
10.1021/bi00346a036
复制
发表时间：
1985-01-01
期刊：
影响因子：
2.9
通讯作者：
RICHARDS, FM
中科院分区：
文献类型：
--
作者：
LEMASTER, DM;RICHARDS, FM
大肠杆菌硫氧还蛋白的10个样品分别同位素富集的残留物类型，通过生长的适当营养缺陷型菌株的培养基上补充一个2 H，15 N-富集的氨基酸。利用95-Hz 1H-15 N酰胺J1偶联对这些样品进行1H双极性-异极性极化实验。从非共振15 N去耦光谱中减去近共振产生的差异图案仅对应于直接键合到15 N核的质子。对于迄今为止观察到的十种不同的富集残基类型，每个标记位置（60）都被观察到为分辨共振。在1H和15 N维度的光谱色散大约为1500 Hz（在500 MHz的场强），而很少明显依赖于残留物类型。除富含甘氨酸的样品外，J1耦合常数的范围并不比测量的精度大得多（1.5-2.0 Hz）。然而，对于甘氨酸残基，J1酰胺偶联值在10 Hz的范围内变化。
Ten samples of Escherichia coli thioredoxin were individually isotopically enriched by residue type via growth of an appropriate auxotrophic strain on media supplemented with one 2H,15N-enriched amino acid. 1H observe-heteronuclear decoupoling experiments were conducted on these samples making use of the 95-Hz 1H-15N amide J1 coupling. Subtraction of near-resonance from off-resonance 15N decoupled spectra generated difference patterns corresponding only to protons directly bonded to 15N nuclei. For the ten different enriched residue types observed to date, every labeled position (60) has been observed as a resolved resonance. The spectral dispersion in both the 1H and the 15N dimensions was roughly 1500 Hz (at 500-MHz field strength) with rather little apparent dependence on residue type. With the exception of the glycine-enriched sample, the range of the J1 coupling constants was not much greater than the precision of the measurements (1.5-2.0 Hz). However, for the glycine residues the J1 amide coupling values varied over a range of 10 Hz.