The Crystal Structure of E. coli rRNA Pseudouridine Synthase RluE

Abstract

Pseudouridine synthase RluE modifies U2457 in a stem of 23 S RNA in Escherichia coli. This modification is located in the peptidyl transferase center of the ribosome. We determined the crystal structures of the C-terminal, catalytic domain of E. coli RluE at 1.2 A resolution and of full-length RluE at 1.6 A resolution. The crystals of the full-length enzyme contain two molecules in the asymmetric unit and in both molecules the N-terminal domain is disordered. The protein has an active site cleft, conserved in all other pseudouridine synthases, that contains invariant Asp and Tyr residues implicated in catalysis. An electropositive surface patch that covers the active site cleft is just wide enough to accommodate an RNA stem. The RNA substrate stem can be docked to this surface such that the catalytic Asp is adjacent to the target base, and a conserved Arg is positioned to help flip the target base out of the stem into the enzyme active site. A flexible RluE specific loop lies close to the conserved region of the stem in the model, and may contribute to substrate specificity. The stem alone is not a good RluE substrate, suggesting RluE makes additional interactions with other regions in the ribosome.

Robust elastic network model: A general modeling for precise understanding of protein dynamics

“…L 7-8 also shows the most significant out-of-plane vibration. These conformational changes would play an important role in RNA binding by inducing the active sites to be closer to the RNA substrate, which is in good agreement with the previous study (Pan et al, 2007).…”

“…As an example, for the case of E. coli RluE (PDB: 2OML) (Pan et al, 2007), RENM shows better B-factor prediction accuracy with lower RMSD and higher CC values than does the basic ENM (Fig. 6A).…”

Steroid Receptor RNA Activator (SRA) Modification by the Human Pseudouridine Synthase 1 (hPus1p): RNA Binding, Activity, and Atomic Model

“…The active site is located in between the two lobes of the catalytic core [6], [17], [18], [19], [20]. PUS enzymes are highly specific, capable of recognising their target uridine when embedded in a particular structural context, avoiding random uridine modification within RNA molecules.…”

Crystal Structure of an RluF–RNA Complex: A Base-Pair Rearrangement Is the Key to Selectivity of RluF for U2604 of the Ribosome

“…A crystal structure of apo TruB with two crystallographically independent protein molecules shows the catalytic domains in distinct conformations that indicate a hinged bending of the central β-sheet, which becomes ordered in the presence of substrate 50. Hinged bending of the central β-sheet is also revealed by crystallographically independent structures of apo RluE51 and is predicted to occur in RluE and RluF based on elastic network model analysis52,53 (Fig. 6c).…”