Development of new 17O NMR spectroscopic techniques for studying biological systems

核磁共振（NMR）光谱是研究分子结构、动力学和化学键合的强大分析技术。过去几十年来，这项技术的发展彻底改变了化学和生物科学。然而，大多数成功的生物系统（如蛋白质和核酸）的NMR研究主要是基于对核自旋量子数为1/2的原子核（如1H、13 C和15 N）的检测。另一方面，氧元素是有机和生物分子中含量最丰富的元素之一，至今还不能用核磁共振波谱法测定。这主要是因为唯一的核磁共振活性氧同位素17 O的核自旋量子数为5/2（称为四极核）。此外，17 O的天然丰度极低，为0.037%。众所周知，四极核很难用核磁共振研究，因为它们经常产生非常宽的核磁共振信号。因此，在生物大分子的背景下，17 O通常被认为是“NMR不可见的”。最近，我们在解决这个具有挑战性的问题的方法上取得了两个突破。首先，我们已经开发了新的NMR方法，允许检测高分辨率的17 O NMR光谱的生物大分子在固态和水溶液中。第二，我们已经证明了一个通用的方法，将17 O纳入重组蛋白与营养缺陷型大肠杆菌。大肠杆菌菌株。在接下来的5年里，我们将把这些新方法应用于两类生物系统：蛋白质和活癌细胞。蛋白质的17 O NMR光谱通常被认为是生物分子NMR的最后前沿。我们将探索17 O NMR在蛋白质结构和动力学研究中所能提供的独特信息。我们将研究使用17 O NMR作为一种新的示踪剂，用于监测葡萄糖和谷氨酰胺代谢在活癌细胞。这将有助于建立17 O NMR作为一种补充工具，可能用于癌症诊断和监测。我们的研究计划的最终目标是使生物分子中发现的所有主要元素（H，C，N和O）都能通过NMR光谱法获得。在这一领域，我们的研究项目一直处于国际领先地位。拟议研究的成果将对化学和生物科学的许多学科产生影响。拟议的研究计划还将为培养下一代高素质人才提供良好的机会，使其具有全球竞争力。

Nuclear magnetic resonance (NMR) spectroscopy is a powerful analytical technique for studying molecular structure, dynamics, and chemical bonding. The development of this technique over the past several decades has revolutionized chemical and biological sciences. However, most successful NMR studies of biological systems such as proteins and nucleic acids are primarily based on detection of atomic nuclei with a nuclear spin quantum number of 1/2 such as 1H, 13C, and 15N. On the other hand, the oxygen element, being one of the most abundant in organic and biological molecules, has not yet been accessible by NMR spectroscopy. This is principally because the only NMR-active oxygen isotope, 17O, has a nuclear spin quantum number of 5/2 (known as a quadrupolar nucleus). In addition, the natural abundance of 17O is extremely low, 0.037%. Quadrupolar nuclei are notoriously difficult to study by NMR because they often give rise to very broad NMR signals. As a result, 17O is very often considered to be "NMR invisible" in the context of biological macromolecules. Recently, we made two breakthroughs on the way of solving this challenging problem. First, we have developed new NMR methods that allow detection of high-resolution 17O NMR spectra for biological macromolecules both in the solid state and in aqueous solution. Second, we have demonstrated a general method of incorporating 17O into recombinant proteins with auxotrophic E. coli strains. In the next 5 years, we will apply these new methods to two types of biological systems: proteins and live cancer cells. 17O NMR spectroscopy of proteins is often considered to be the last frontier of biomolecular NMR. We will explore the unique information that 17O NMR can offer in the study of protein structure and dynamics. We will investigate the use of 17O NMR as a new tracer for monitoring glucose and glutamine metabolism in live cancer cells. This will help establish 17O NMR as a complementary tool potentially useful for cancer diagnosis and monitoring. The ultimate goal of our research program is to make all major elements (H, C, N, and O) found in biological molecules equally accessible by NMR spectroscopy. In this field, our research program has been an international leader. The outcomes from the proposed research will have impact in many disciplines in chemical and biological sciences. The proposed research program will also provide excellent opportunities for training of the next generation of highly qualified personnel to be competitive globally.