## 3 Structural Basis of Transcription Elongation  
Crystals of the transcription elongation complex were prepared using _T. thermophilus_ RNAP and a synthetic DNA/RNA scaffold, and structures were determined with and without a nucleotide triphosphate substrate (PDB: 2O5I, 2O5J) [42,43]. Structures revealed atomic details of RNAP and DNA/RNA interactions within the DNA binding main channel and the RNA exit channel and showed how the RNAP mobile element trigger loop changes conformation throughout nucleotide substrate selection and phosphodiester bond formation. During RNA extension, RNAP temporarily stops transcription (transcription pausing) by several mechanisms including \(\sigma\) factor-dependent promoter proximal pausing, elemental pausing, RNA hairpin-dependent pausing and backtrack pausing (Figure 1) [52]. The structure of an elemental-paused elongation complex using _Thermus_ RNAP and DNA/RNA scaffolds derived from the _E. coli_ his-pause sequence showed a unique RNAP conformation including an open-clamp and kinked bridge helix that may inhibit processive RNA extension (PDB: 4 GZY, 4 GZZ) [44]. Misincorporated nucleotides in RNA transcripts slow down the rate of transcription and result in the backward movement of RNAP (backtracking), which ejects the 3' RNA end into the secondary channel and later cleaves misincorporated nucleotides by RNAP endonuclease activity (Figure 1). This activity is further stimulated by the elongation factor GreA. The structure of this single-nucleotide-backtracked elongation complex showed a sharply bent RNA backbone at the unpaired RNA base, which accommodated the proofreading cavity of the RNAP active site (PDB: 4 WQS) [45]. This study also revealed the structure of RNAP in complex with GreA-like protein, showing that the GreA coiled-coil domain containing acidic residues binds to the RNAP active site through the secondary channel and coordinates Mg\({}^{2+}\), which is essential for RNA cleavage (PDB: 4 WQT). The transcription factor RapA is an ATP-dependent DNA translocase, which reactivates the stalled elongation complex (Figure 1). The structure of the elongation complex with RapA showed that RapA binds around the RNA exit site, making the RNA channel just wide enough for single-stranded RNA to pass (PDB: 4S20) [53]. Promoter-Dependent Transcription: How RNA Polymerase Recognizes Promoter DNA Sequences and Initiates Transcription  
Bacterial RNAP recognizes promoter DNA and initiates transcription as the holoenzyme form containing the catalytic core enzyme plus the promoter recognition \(\sigma\) factor (Figure 1). Structures of the holoenzyme containing the group I \(\sigma\) factor \(\sigma^{\rm A}\) (also known as SigA), isolated from _T. aquaticus_ and _T. thermophilus_, showed that three \(\sigma\) domains (\(\sigma\) domains 2, 3, and 4) arranged on the surface of the core enzyme whose function was to recognize the \(-\)35 and\(-\)10 elements (separated by \(\sim\)17 bp DNA) (PDB: 1L9U, 1 IW7) [32, 33] (Table 1). The structure of the holoenzyme in complex with fork junction promoter DNA (\(-\)41 to \(-\)7 relative to the transcription start site at \(+\)1) showed that the promoter DNA lies across one face of the holoenzyme and is positioned completely outside the RNAP cleft (PDB: 1L9Z) [38]. Universally conserved aromatic residues in the \(\sigma\) region 2.3 are ideally positioned to stack on the exposed face of the base pair at the upstream edge of the transcription bubble. Atomic details of \(-\)35 element recognition by \(\sigma\) domain 4 (\(\sigma\)4), as revealed by co-crystal structure, showed that the helix-turn-helix motif of \(\sigma\)4 reads the hexameric DNA sequence (PDB: 1 KU7) [16]. Details of the interaction between \(\sigma\)2 and the \(-\)10 element were obtained from the structure of \(\sigma\)2 bound to the single-stranded \(-\)10 element (PDB: 3 UGO, 3 UGP) [18] and the _T. thermophilus_ RNAP holoenzyme in complex with the promoter DNA fragment (\(-\)12 to \(+\)12 on DNA) (PDB: 4G7H, 4G7O) [39].