High-resolution measurement of long-range distances in RNA: pulse EPR spectroscopy with TEMPO-labeled nucleotides
نویسندگان
چکیده
SI1: Sample characterization by HPLC and CW-EPR spectroscopy. Figure S1: (a) Anion exchange traces of purified single-stranded double-C T -labeled RNAs. (b) Spin concentration calibration curve. (c) RNA labelling efficiency. SI2: Examination of orientation selection. Figure S2: Highand low-power Q-band DEER on [13,28] RNA duplex. Figure S3: High-power Q-band DEER on different RNA samples. Figure S4: PELDOR/DEER traces with longer evolution time. SI3: Effect of background subtraction on distances and distributions. Figure S5: Analysis with four different background corrections of samples 1, 3, 6 and 7.
منابع مشابه
Site-specific incorporation of nitroxide spin-labels into internal sites of the TAR RNA; structure-dependent dynamics of RNA by EPR spectroscopy.
RNA can catalyze chemical reactions1 and interacts in specific ways with other macromolecules, such as proteins. The mechanistic understanding of RNA function relies on structural information; however, solving crystal structures of complex RNA molecules remains challenging.2 Therefore, other biophysical techniques have been used to determine the position of RNA secondary structure elements (i.e...
متن کاملComplementary-addressed site-directed spin labeling of long natural RNAs
Nanoscale distance measurements by pulse dipolar Electron paramagnetic resonance (EPR) spectroscopy allow new insights into the structure and dynamics of complex biopolymers. EPR detection requires site directed spin labeling (SDSL) of biomolecule(s), which remained challenging for long RNAs up-to-date. Here, we demonstrate that novel complementary-addressed SDSL approach allows efficient spin ...
متن کاملDemystifying EPR: A Rookie Guide to the Application of Electron Paramagnetic Resonance Spectroscopy on Biomolecules
Electron Paramagnetic Resonance (EPR) spectroscopy, also known as Electron Spin Resonance(ESR) especially among physicists, is a strong and versatile spectroscopic method forinvestigation of paramagnetic systems, i.e. systems like free radicals and most transition metalions, which have unpaired electrons. The sensitivity and selectivity of EPR are notable andintriguing as compared to other spec...
متن کاملNanometer-Range Distance Measurement in a Protein Using Mn Tags
Pulse electron paramagnetic resonance measurements of long-range (nm scale) distances between spin labels site-specifically attached to biomacromolecules have proven highly effective in structural studies. The most commonly used spin labels are stable nitroxide radicals, and measurements are usually carried out at X-band frequencies (∼9.5 GHz, 0.35 T). Higher magnetic fields open new possibilit...
متن کاملProbing secondary structures of spin-labeled RNA by pulsed EPR spectroscopy.
The ability of RNA to interconvert between multiple conformational states is essential for the diversity of biological functions that have been discovered in the recent past. For example, the correct operation of regulatory RNA elements, such as riboswitches, is based on the precise interplay of alternative RNA conformations. Studying the molecular mechanisms of RNA function entails probing RNA...
متن کامل