A TROSY CPMG sequence for characterizing chemical exchange in large proteins
A TROSY CPMG sequence for characterizing chemical exchange in large proteins
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DOI：
10.1023/a:1008355631073
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发表时间：
1999-10-01
影响因子：
2.7
通讯作者：
Palmer, AG
中科院分区：
文献类型：
--
作者：
Loria, JP;Rance, M;Palmer, AG
本文介绍了一种新的核磁共振自旋弛豫实验，用于测量N-15标记的大分子中约0.5 ~ 5 ms的化学交换时间常数。脉冲序列基于Carr-Purcell-Meiboom-Gill技术[卡尔和珀塞尔（1954）物理评论，94，630-638; Meiboom和Gill（1958）Rev. Sci.仪器：29，688-691; Loria等人（1999）J. Am.化学会，121，2331-2332]，但实施TROSY选择[Pervushin等人（1997）Proc.Natl. Acad. Sci. USA，94，12366-12371]，以允许测量对H-1-N-15标量偶联二重态的较窄组分的交换谱线增宽贡献。这种修改扩展了由于在较大的大分子中的横向磁化强度的快速衰减而对弛豫测量施加的尺寸限制。在碱性胰蛋白酶抑制剂的[U-98%N-15]标记样品和磷酸丙糖异构酶（一种54 kDa同源二聚体蛋白）的[U-83%H-2，U-98%N-15]标记样品上证明了新的TROSY-CPMG实验。
A new NMR spin relaxation experiment is described for measuring chemical exchange time constants from approximately 0.5 ms to 5 ms in N-15-labeled macromolecules. The pulse sequence is based on the Carr-Purcell-Meiboom-Gill technique [Carr and Purcell (1954) Phys. Rev., 94, 630-638; Meiboom and Gill (1958) Rev. Sci. Instrum., 29, 688-691; Loria et al. (1999) J. Am. Chem. Soc., 121, 2331-2332], but implements TROSY selection [Pervushin et al. (1997) Proc. Natl. Acad. Sci. USA, 94, 12366-12371] to permit measurement of exchange linebroadening contributions to the narrower component of the H-1-N-15 scalar-coupled doublet. This modification extends the size limitation imposed on relaxation measurements due to the fast decay of transverse magnetization in larger macromolecules. The new TROSY-CPMG experiment is demonstrated on a [U-98% N-15] labeled sample of basic pancreatic trypsin inhibitor and a [U-83% H-2, U-98% N-15] labeled sample of triosephosphate isomerase, a 54 kDa homodimeric protein.