Lundström Patrik, Hansen D Flemming, Kay Lewis E
Department of Medical Genetics, The University of Toronto, Toronto, ON, Canada, M5S 1A8.
J Biomol NMR. 2008 Sep;42(1):35-47. doi: 10.1007/s10858-008-9260-4. Epub 2008 Sep 2.
Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy has emerged as a powerful method for quantifying chemical shifts of excited protein states. For many applications of the technique that involve the measurement of relaxation rates of carbon magnetization it is necessary to prepare samples with isolated (13)C spins so that experiments do not suffer from magnetization transfer between coupled carbon spins that would otherwise occur during the CPMG pulse train. In the case of (13)CO experiments however the large separation between (13)CO and (13)C(alpha) chemical shifts offers hope that robust (13)CO dispersion profiles can be recorded on uniformly (13)C labeled samples, leading to the extraction of accurate (13)CO chemical shifts of the invisible, excited state. Here we compare such chemical shifts recorded on samples that are selectively labeled, prepared using [1-(13)C]-pyruvate and NaH(13)CO(3,) or uniformly labeled, generated from (13)C-glucose. Very similar (13)CO chemical shifts are obtained from analysis of CPMG experiments recorded on both samples, and comparison with chemical shifts measured using a second approach establishes that the shifts measured from relaxation dispersion are very accurate.
卡尔-珀塞尔-梅博姆-吉尔（CPMG）弛豫色散核磁共振（NMR）光谱已成为一种用于量化激发态蛋白质化学位移的强大方法。对于该技术的许多应用，涉及碳磁化弛豫率的测量，有必要制备具有孤立（13）C自旋的样品，以便实验不会受到耦合碳自旋之间的磁化转移影响，否则在CPMG脉冲序列期间会发生这种转移。然而，在（13）CO实验的情况下，（13）CO和（13）Cα化学位移之间的大间隔提供了一种希望，即可以在均匀（13）C标记的样品上记录稳健的（13）CO色散谱，从而提取不可见激发态的准确（13）CO化学位移。在这里，我们比较了在使用[1-（13）C]-丙酮酸和NaH（13）CO3制备的选择性标记样品或由（13）C-葡萄糖产生的均匀标记样品上记录的此类化学位移。对在两个样品上记录的CPMG实验进行分析，得到了非常相似的（13）CO化学位移，并且与使用第二种方法测量的化学位移进行比较，确定从弛豫色散测量的位移非常准确。
