A new isoindoline-derived benzimidazole nitroxide spin label, ^ImUm, was synthesized and incorporated into RNA oligoribonucleotides. ^ImUm is the first example of a conformationally unambiguous spin label for RNA, in which the nitroxide N–O bond lies on the same axis as the single bond used to attach the rigid isoindoline-based spin label to a uridine base. This results in minimal displacement of the nitroxide upon rotation of this single bond, which is a useful property for a label to be used for distance measurements. Continuous-wave (CW) EPR measurements of RNA duplexes containing ^ImUm indicate a restricted rotation around this single bond, presumably due to an intramolecular hydrogen bond between the benzimidazole N–H and O4 of the uracil. Orientation-selective pulsed electron–electron double resonance (PELDOR, also called double electron–electron resonance, or DEER) distance measurements between two spin labels in two RNA duplexes showed in one case a strong orientation dependence, further confirming the restricted motion of the spin labels in RNA duplexes.

中文翻译：

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半刚性的异吲哚啉衍生的一氧化氮自旋标记，用于RNA †

合成了新的异吲哚啉衍生的苯并咪唑一氧化氮自旋标记^Im Um，并将其掺入RNA寡核糖核苷酸中。^Im Um是RNA构象明确的自旋标记的第一个例子，其中的氮氧化物N–O键与用于将基于刚性二氢吲哚啉的自旋标记附着于尿苷碱基的单键位于同一轴上。这导致该单键旋转时氮氧化物的位移最小，这对于用于距离测量的标签是有用的属性。含^Im Um的RNA双链体的连续波（CW）EPR测量表示围绕该单键的旋转受限，可能是由于苯并咪唑N–H和尿嘧啶O4之间存在分子内氢键。两种RNA双链体中两个自旋标记之间的方向选择脉冲电子-电子双共振（PELDOR，也称为双电子-电子共振或DEER）距离测量在一种情况下显示出强烈的方向依赖性，进一步证实了自旋运动受限制RNA双链体中的标记。