Fig. 5: Cryo-EM structure of product states of the RT-T/P-dATP complex.

Location of the 2′-O-methyl moiety (2′-OMe) in states a R-1, b P_2min, and c P_6min of the RT-T/P-dATP complex. a In states R-1, R-2, and T_8sec (R-2 and T_8sec not displayed for clarity purposes) density (yellow mesh) for the 2′-OMe (Cm, red sticks) is observed in position +2 and +4 and is clearly absent in positions +3 and +5 of the template strand (map displayed with the sharpening of B-factor = −60 Ų contoured at 12σ). b In-state P_2min density for 2′-OMe is observed in positions +2, +3, +4, and +5 as a mixture of translocated and non-translocated molecules. *Cm denotes partial occupancy of the 2′-OMe template scaffold. Map density is colored with yellow mesh at position +4, and green mesh at positions +3 and +5 (contoured at 7.5σ and sharpened at B-factor of −40 Ų). c In P_6min density for 2′-OMe (green mesh) is observed only in positions +3 and +5, indicating all molecules have translocated one position (map displayed with B-factor sharpening = −40 Ų contoured at 8.5σ). d, e Ball-and-stick representation of the active site in state P_6min with a mixture of d products (added dAMP and pyrophosphate (PPi)) and e reactants (i.e., dATP). f Overlay of density maps of states R-1 and P_6min of the RT-T/P-dATP complex. Black arrows point to the growth of density for the newly formed phosphodiester bond; red arrows indicate thinning of density around O3A; green arrows point to stretched density for the leaving pyrophosphate (PPi) group (densities contoured at σ = 12.5 and σ = 9.5, for R-1 and P_6min, respectively. Maps are sharpened at B-factor of −60 Ų). g Detail of density maps inside box in (f), displayed at high threshold (σ = 35 and σ = 14, for state R-1 and state P_6min, respectively. B-factor sharpening = −100 Ų). h The PPi is stabilized by metal B and surrounding amino acids K65, D113, and K220 (Fig. 6c). Distances are indicated next to dashed lines (in angstroms).