13C LABELED GLYCEROL

13C 标记甘油

• 批准号: 6309047
• 负责人: DAVID LEMASTER
• 金额: $ 2.74万
• 依托单位: UNIVERSITY OF CALIF-LOS ALAMOS NAT LAB
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-01 至 2002-01-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6309047
• 关键词: bacteria; biomedical resource; nuclear magnetic resonance spectroscopy; proteins

The SIR provided [13C, 1,3-'H4 ]Glycerol; 14g Uniform 13C enrichment has become widely used for resonance assignment studies of proteins and, more recently, nucleic acids. Along with the various benefits, there are several limitations to this labeling pattern which stem from the presence of one bond 13C-13C couplings. In addition to the degraded reslution in the 13C dimension which can only partially be alleviated by constant time techniques, no inderct detected 13C relaxation experiments have been developed which cercumvent the artifacts due to the 13C-13C scalar interactions. The alternating carbon enrichment procedure we have developed for E. coli expression systems allows for both relaxation experiments as well as various 13C editing assignment experiments. This method involves culturing the organisms using [2-13C]- or [1,3-13C21glycerol as a carbon source. Furthermore, we have demonstrated that by combining random fractional deuteration with selective 13C enrichment, one can analyze lH detected relaxation of methylene carbons which has heretofore not been feasible. As a result by using the two opposite patterns of 13C enrichment it should be possible to deterine the motional parameters of nearly all sites of a protein or nucleic acid molecule. In addition, particularly in the case of nucleic acids, we anticipate that this approach to selective labeling will offer a substantial complement to structural studies via uniform 13C enrichment.

SIR提供[13 C，1，3-'H4]甘油; 14 g均匀13 C 富集已经广泛用于共振分配研究， 蛋白质和最近的核酸。 沿着各种 但是，这种标记模式有几个限制， 源于一个键13 C-13 C偶联的存在。 除了 在13 C维度上的退化结果，其仅可以部分地 通过恒定时间技术缓解，未检测到13 C 松弛实验已经开发出， 由于13 C-13 C标量相互作用而产生的伪影。 交替 碳富集程序，我们已经开发了E。杆菌表达 系统允许弛豫实验以及各种13 C 编辑作业实验。 该方法包括培养 使用[2- 13 C]-或[1，3 - 13 C 21]甘油作为碳源的生物。 此外，我们已经证明，通过结合随机分数 使用选择性13 C富集的氘代，可以分析检测到的1H 亚甲基碳的弛豫，迄今为止还没有被 可行 因此，通过使用13 C的两种相反模式， 富集，应该可以确定运动参数 蛋白质或核酸分子的几乎所有位点。 在 此外，特别是在核酸的情况下，我们预计 这种选择性标记的方法将提供一个实质性的 通过均匀的13 C富集补充结构研究。