Two-dimensional measurement of proton T1rho relaxation in unlabeled proteins: mobility changes in alpha-bungarotoxin upon binding of an acetylcholine receptor peptide.
Two-dimensional measurement of proton T1rho relaxation in unlabeled proteins: mobility changes in alpha-bungarotoxin upon binding of an acetylcholine receptor peptide.
复制标题
未标记蛋白质中质子 T1rho 弛豫的二维测量：结合乙酰胆碱受体肽后 α-金环蛇毒素的迁移率变化。
DOI：
10.1021/bi050645h
复制
发表时间：
2005
期刊：
影响因子：
2.9
通讯作者：
Anglister,Jacob
中科院分区：
文献类型：
--
作者：
Samson,AbrahamO;Chill,JordanH;Anglister,Jacob
描述了一种测量未标记蛋白质质子T1 ρ弛豫时间的方法，该方法在HOHAHA脉冲序列的初始非选择性90°激发后，使用可变自旋锁脉冲。本实验应用于α-银环蛇毒素（α-BTX）及其与乙酰胆碱受体（AChR）α亚基衍生的25个残基的肽的复合物。高T1 ρ值和局部运动增加之间有很好的相关性。在游离形式中，根据α-BTX与AChR肽复合物的NMR结构和α-BTX与AChR的模型，与受体结合相关的毒素残基[Samson，A. O.，等人（2002）Neuron 35，319 - 332]显示高迁移率。当AChR肽结合时，表现出参与受体α亚基结合的残基的弛豫时间和运动水平的降低，而参与结合γ和δ亚基的残基保持其运动性。此外，定量T1 ρ测量使我们能够证实与毒素强烈相互作用的AChR决定簇的边界的映射[Samson，A. O.，等人（2001）Biochemistry 40，5464 - 5473]，并且可以类似地应用于其中肽代表两种相互作用蛋白质之一的其他蛋白质复合物。所提出的方法是有利的，因为它的简单性，通用性和时间效率，并铺平了道路，为未来的调查质子弛豫速率在小的未标记的蛋白质。
A method for the measurement of protonT1ρrelaxation times in unlabeled proteins is described using a variable spin-lock pulse after the initial nonselective 90° excitation in a HOHAHA pulse sequence. The experiment is applied to α-bungarotoxin (α-BTX) and its complex with a 25-residue peptide derived from the acetylcholine receptor (AChR) α-subunit. A good correlation between highT1ρvalues and increased local motion is revealed. In the free form, toxin residues associated with receptor binding according to the NMR structure of the α-BTX complex with an AChR peptide and the model for α-BTX with the AChR [Samson, A. O., et al. (2002)Neuron35, 319−332] display high mobility. When the AChR peptide binds, a decrease in the relaxation times and the level of motion of residues involved in binding of the receptor α-subunit is exhibited, while residues implicated in binding γ- and δ-subunits retain their mobility. In addition, the quantitativeT1ρmeasurements enable us to corroborate the mapping of boundaries of the AChR determinant strongly interacting with the toxin [Samson, A. O., et al. (2001)Biochemistry40, 5464−5473] and can similarly be applied to other protein complexes in which peptides represent one of the two interacting proteins. The presented method is advantageous because of its simplicity, generality, and time efficiency and paves the way for future investigation of proton relaxation rates in small unlabeled proteins.