Liu A, Hu W, Qamar S, Majumdar A
Cellular Biochemistry & Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
J Biomol NMR. 2000 May;17(1):55-61. doi: 10.1023/a:1008340116418.
In this paper, we demonstrate that the sensitivity of triple-resonance NMR experiments can be enhanced significantly through quenching scalar coupling mediated relaxation by using composite-pulse decoupling (CPD) or.an adiabatic decoupling sequence on aliphatic, in particular alpha-carbons in 13C/15N-labeled proteins. The CPD-HNCO experiment renders 50% sensitivity enhancement over the conventional CT-HNCO experiment performed on a 12 kDa FK506 binding protein, when a total of 266 ms of amide nitrogen-carbonyl carbon defocusing and refocusing periods is employed. This is a typical time period for the direct detection of hydrogen bonds in proteins via trans-hydrogen bond 3hJ(NC') couplings. The experimental data fit theoretical analysis well. The significant enhancement in sensitivity makes the experiment more applicable to larger-sized proteins without resorting to perdeuteration.
在本文中，我们证明了通过使用复合脉冲去耦（CPD）或绝热去耦序列淬灭标量耦合介导的弛豫，可以显著提高三共振核磁共振实验的灵敏度，该实验针对13C/15N标记蛋白质中的脂肪族，特别是α-碳原子。当总共使用266 ms的酰胺氮-羰基碳散焦和重聚焦周期时，CPD-HNCO实验相对于在12 kDa FK506结合蛋白上进行的传统CT-HNCO实验，灵敏度提高了50%。这是通过反式氢键3hJ(NC')耦合直接检测蛋白质中氢键的典型时间段。实验数据与理论分析吻合良好。灵敏度的显著提高使得该实验更适用于更大尺寸的蛋白质，而无需采用全氘代。
