Catalytic mechanism of Ire1 RNase. (a) Catalytic residues of RNase T1 and RNase A with productively bound nucleotides (PDB ID 1r5c and 1rga). (b) Left panel: simulated annealing (1000 K) omit-electron density map Fo-Fc for the helix-loop element (HLE; green) and adjacent loop containing Y1043 and the helix α4 of RNase domain (contoured at 5σ). Refinement was conducted with residues 1032-1057 deleted from all 7 monomers in the asymmetric unit, without using non-crystallographic symmetry (NCS). An alternative view of the electron density (after rotation around vertical axis) is shown in Additional file 1, Figure S4. Right panel: position of the scissile phosphate in the active site of Ire1 RNase. The strongest peak of the electron density (Fo-Fc, contoured at 12σ) was used to position the phosphate. Side chains were taken from PDB ID 3fbv). (c) Two plausible catalytic mechanisms in the active site of Ire1 RNase. (d) Enzyme titration profiles for Ire1KR32, Ire1KR32(Y1043F) and for Ire1KR32(R1039A) under single-turnover conditions. Reactions were conducted as in Figure 1c.
Korennykh et al. BMC Biology 2011 9:47 doi:10.1186/1741-7007-9-47