Figure 1. Protected imino protons indicate secondary structure formation. (a) Sequence and secondary structure of the apical R/G stem–loop RNA. (b) Imino proton region of the one‐dimensional spectra of the 27mer, apical R/G stem–loop RNA, recorded with a 600‐MHz spectrometer at 278 K. The assignments are annotated for the imino proton resonances [ 9].

Figure 2. Assessment of resonance dispersion in natural abundance – HSQC spectrum. (a) A zoomed aromatic region of the natural abundance – HSQC spectrum of the apical R/G stem–loop RNA at 600 MHz, recorded at 303 K. The assignments of the C6/C8–H6/H8, C2–H2, and C5–H5 correlations are indicated. The C5–H5 peaks of pyrimidines are folded from their typical chemical shift range of 90–110 ppm. (b) Sequence and secondary structure of the apical R/G stem–loop RNA [ 9].

Figure 3. NOE connectivities of imino, amino, and aromatic protons in the GC (A) and AU (b) base pairs.

Figure 4. One‐bond (a), two‐bond (b), and long‐range (c) spin–spin scalar couplings in RNA bases.

Figure 5. The coherence transfer pathways for the H8–C8–N9–C1′–H1′ (a) and H6–C6–N1–C1′–H1′ (b) correlations in purine and pyrimidine nucleotides, respectively.

Figure 6. “Base–sugar sequential walk” in the two‐dimensional – NOESY spectra of an A‐form RNA double helix. A close‐up of the two‐dimensional NOE spectrum of the apical R/G stem–loop RNA, recorded with a 600‐MHz spectrometer (mixing time of 300 ms) at 303 K in D ₂O. The assignments for the intra‐ and inter‐residue connectivities are shown.

Figure 7. NOE sequential connectivities in an A‐RNA helix.

Figure 8. Two‐bond and three‐bond spin–spin scalar couplings used in HCP experiments to establish the inter‐residual through‐bond connectivities.

Figure 9. Flowchart of sample preparation.

Figure 10. Flowchart of experimental data collection.

Figure 11. Flowchart of structure calculation.

Figure 12. T7 class III promoter region and the resulting transcript. (a) T7 class III promoter region with indicated binding and initiating regions. Position +1 indicates the first residue of the transcript. (b) The RNA transcript is generated from position +1 of the template strand. The recommended nucleotides for positions +1 and +2 are shown. These positions are strongly conserved within the initiating domain across various bacteriophage promoters (class III T7, T3 and SP6, and T7 f2.5 class II).