New trityl radicals for studies of biopolymers by NMR and EPR spectroscopies

核磁共振（NMR）和电子顺磁共振（EPR）光谱是研究生物聚合物的常用方法。核磁共振可以提供高分辨率的结构，但它本身是一种不敏感的技术，这限制了它的应用。然而，快速兴起的动态核极化（DNP）领域通过将自旋极化从电子转移到核自旋，导致核磁共振的灵敏度提高了数百倍。本提案的主要目的是为DNP-NMR制备新的基于三芳基甲基（三烷基）的单自由基和双自由基。螺环己醇基衍生的三烷基自由基将解决三烷基水溶性有限及其电子弛豫速率的缺点。除了利用单自由基解决三苯基的水溶性外，还将制备新的双自由基，特别是氮氧基-三苯基自由基。此外，还制备了两类新的三基基自由基：三基基-三基基双自由基和1,3-二苯基-2-苯丙基(BDPA)-三基基双自由基。这些窄线双基将用于几种低γ核（13C， 15N和29Si）的极化增加。由于三烷基自由基在还原条件下具有很高的稳定性，一些新的自由基将用于细胞内DNP-NMR。马来酰亚胺和碘乙酰胺衍生的三烷基自由基将用于蛋白质的定向自旋标记，后者也将用于核酸，用于EPR光谱的研究。三基自由基也会附着在荧光标记的细胞穿透肽上，以供细胞递送。这种肽缀合物将使荧光显微镜能够可视化三烷基自由基的细胞位置。选定的三烷基自由基也将用于最先进的脉冲DNP-NMR实验。所提出的自由基具有显著影响DNP-NMR和EPR领域的潜力。

Nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopies are commonly used for investigation of biopolymers. NMR can provide high-resolution structures but is an inherently insensitive technique, which limits its applications. However, the rapidly emerging field of dynamic nuclear polarization (DNP) has led to an increased sensitivity of NMR, of up to several hundred-fold, by transferring spin polarization from electrons to nuclear spins. The main goal of this proposal is the preparation of new triarylmethyl (trityl)-based mono- as well as biradicals for DNP-NMR. A spirocyclohexanolyl-derived trityl radical will address shortcomings of limited water-solubility of trityls and of their electron relaxation rates. In addition to addressing water-solubility of trityls for biological applications with monoradicals, new biradicals will also be prepared, in particular nitroxide-trityl radicals. Moreover, two new classes of trityl-based radicals will be prepared, trityl-trityl biradicals and 1,3-bisdiphenylene-2-phenylallyl (BDPA)-trityl biradicals. These narrow-line biradicals will be prepared for increased polarization of several low-gamma nuclei (13C, 15N and 29Si). Since trityl radicals possess high stability under reducing conditions, some of the new radicals will be used for in-cell DNP-NMR. Maleimide- and iodoacetamide-derived trityl radicals will be prepared for site-directed spin labeling of proteins and the latter will also be applied for nucleic acids, for studies by EPR spectroscopy. Trityl radicals will also be attached to a fluorescently-labeled, cell-penetrating peptide for cellular delivery. Such peptide conjugates will enable visualization of the cellular location of the trityl radicals by fluorescence microscopy. Selected trityl radicals will also be used for state-of-the-art pulsed DNP-NMR experiments. The proposed radicals have the potential to significantly impact the field of DNP-NMR, as well as EPR.