Fig. 4: Composite model of the arKEOPS holo-enzyme bound to tRNA is suggestive of an enzyme–substrate complex.
From: A substrate binding model for the KEOPS tRNA modifying complex
a Stereo surface representation of arKEOPS bound to mjtRNALys (see methods for model construction). Black ribbon represents the position of tRNA prior to energy minimization. Note the severe steric clash between tRNA and the Kae1 subunit. Red ribbon represents the position of tRNA following energy minimization (see Methods). The steric clash between tRNA and Kae1 is relieved by a 21° rigid body pivot of tRNA centered at the junction between tRNA nucleotides G73 and C74. For clarity, only the positions of the last four bases of the tRNA are shown. b Representative two-dimensional class averages of KEOPS with and without tRNA determined by single particle negative stain electron microscopy. Additional density observed in the tRNA-bound specimen is indicated by the white triangles. c Three-dimensional reconstructions of KEOPS with and without tRNA derived from the two-dimensional-classes. (Top) atomic models and their superimposed theoretical envelopes of KEOPS alone (left) and KEOPS bound to tRNA (right). (Bottom) atomic models fitted into the 3D reconstructions of KEOPS alone (left) and KEOPS bound to tRNA (right). d Zoom-in stereo view of the active site region of mjKae1 with the KEOPS–tRNA model (blue) highlighting the distance between the modeled position of the A37 modification acceptor site and the modification acceptor site of tobramycin as well as the predicted distances to the position of the donor atom of TC-AMP. The TobZ-tobramycin enzyme–substrate complex (PDB 3VET) was superimposed on the Kae1 subunit in the composite model shown in (a) by alignment of metal binding residues using Pymol.
