Water uptake in vesicles and the subsequent exchange between water protons and amide –NH protons in amino acids can be followed by a new, highly sensitive, type of magnetic resonance spectroscopy: dynamic nuclear polarisation (DNP)-enhanced NMR in the liquid state. Water hydrogen atoms are detected prior to and after their transfer to molecular sites in peptides and proteins featuring highly-accessible proton-exchangeable groups, as is the case for the –NH groups of intrinsically disordered proteins. The detected rates for amide proton–water proton exchange can be modulated by membrane-crossing rates, when a membrane channel is interposed. We hyperpolarised water proton spins via dynamic nuclear polarisation followed by sample dissolution (d-DNP) and transferred the created polarisation to –NH groups with high solvent accessibility in an intrinsically disordered protein domain. This domain is the membrane anchor of c-Src kinase, whose activity controls cell proliferation. The hindrance of effective water proton transfer rate constants observed in free solvent when a membrane-crossing step is involved is discussed. This study aims to assess the feasibility of recently-introduced hyperpolarised (DNP-enhanced) NMR to assess water membrane crossing dynamics.

中文翻译：

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超极化NMR通过与生物分子交换来跟踪水质子通过膜通道的传输

囊泡中的水分吸收以及随后的水质子与氨基酸中的酰胺-NH质子之间的交换之后，可以得到一种新型的，高灵敏度的磁共振波谱：动态核极化（DNP）增强的液态NMR。在氢和氢原子转移到具有高度可及质子交换基团的肽和蛋白质的分子部位之前和之后，都会检测到氢原子，就像固有杂乱的蛋白质的–NH基团一样。当插入膜通道时，可以通过膜交叉速率来调节检测到的酰胺质子-水质交换的速率。我们通过将超极化水质子自旋动态核极化，然后进行样品溶解（d-DNP），并将产生的极化转移至内在无序蛋白质结构域中具有高溶剂可及性的–NH基团。该结构域是c-Src激酶的膜锚，其活性控制细胞增殖。讨论了涉及跨膜步骤时在游离溶剂中观察到的有效水质子转移速率常数的障碍。这项研究旨在评估最近引入的超极化（DNP增强）NMR评估水膜穿越动力学的可行性。