Figure 1. CryoEM structure of the Eco 6S RNA/Eσ⁷⁰ complex.

(A) (top) Secondary structure of wild-type Eco 6S RNA as observed in the cryoEM structure. Structural elements of the 6S RNA are labeled (CS, closing stem; DB, downstream bulge; DD, downstream duplex; CB, central bubble; UD1-3, upstream duplexes 1-3; UB1-2, upstream bulges 1-2; UTL, upstream terminal loop). The sequence is color-coded according to the RNase footprinting and localized hydroxyl-radical cleavage results comparing 6S RNA with and without Eσ⁷⁰ (Figure S1B). RNA positions protected from RNase V1 and/or RNase A cleavage in the presence of Eσ⁷⁰ are colored green. RNA positions showing hypersensitivity to RNAse V1 in the presence of Eσ⁷⁰ are colored red. RNA positions efficiently cleaved by hydroxyl radicals generated from Fe²⁺ in place of the RNAP active site Mg²⁺ are boxed in yellow (Wassarman and Saecker, 2006). The position of pRNA synthesis initiation (U44) is indicated by a bent arrow.

(below) Secondary structure of 6S RNA* used in cryoEM structure determination, generated by truncating the CS and shuffling the sequence of the DD to give DD* (shuffled sequences highlighted in the orange box).

(B) The 3.8 Å resolution cryoEM density map of Eco 6S RNA/Eσ⁷⁰ is rendered as a transparent surface colored as shown. Superimposed is the final refined model; the RNAP is shown as a backbone ribbon, the 6S RNA is shown in stick format.

(C) The 3.8-Å resolution cryoEM density map with the superimposed model of only the 6S RNA. Shown for reference is the RNAP active site Mg²⁺ ion (yellow sphere).

(D) Shown is just the promoter DNA after superimposing the Eσ⁷⁰ from the crystal structure of an open promoter complex (PDB ID 4XLN) (Bae et al., 2015) onto the 6S RNA/Eσ⁷⁰ structure. The promoter DNA is colored blue but the -35 and -10 elements of the DNA are colored yellow. The -6G of the discriminator and the Core Recognition Element (CRE) are colored violet. Shown for reference is the RNAP active site Mg²⁺ ion (yellow sphere).

See also Table S1 and Figures S1 – S4.