Figure 3.

Structural characterization of GRTH. (A) Domain 1 of DDX25 showing the binding sites of ATP and RNA along with the DEAD-box sequence and the relative positions of T239 and R242. Both residues are solvent exposed and located at the GRTH/PKA binding interface. (B) GRTH/PKA interface on GRTH indicating the basic (blue) and acidic (red) residues surrounding T239; most of these residues are unique to DDX25 including T239 itself; only 5 out of the 12 residues shown are common between DDX25 and DDX19. (C) Intramolecular H-bond network of residues at the GRTH/PKA interface for wild-type GRTH. The network was identified from MD simulations and deemed statistically significant in frequency: red lines indicate strong/persistent H-bonds; blue lines are either weak bonds or strong bonds developed upon mutations of interfacial residues; simulations were carried out for single mutants E165A, K234A, K234D, K240A, K240D, D237H, D237A, R242H (neutral and protonated histidine), R242A and for the double mutants E165A + E272A, E165A + R242A, E165A + K240A. The MD results were used to suggest mutations to be carried out experimentally. (D) like in (C) but for H242 (protonated) observed in infertile patients¹². (E) Surface electrostatic potential on GRTH at the GRTH/PKA interface; same orientation as in (B) (red: negative field, blue: positive field, white: neutral field; drawn at the same magnitude of positive and negative field intensities). Note the negative elongated crevice flanked by two regions of positive field; hydrophobic regions are generally outside the interface. (F) Details of surface topography at the interface with T239 highlighted in green.