Cocrystal structure of an RNA sulfuration enzyme MnmA and tRNA-Glu in the initial tRNA binding state
Classification:
Transferase/RNA
Technique:
X-Ray Diffraction
Resolution:
3.1
R value free:
0.269
R value observed:
0.219
R value work:
0.219
Abstract of the PDB Structure's related Publication:
Uridine at the first anticodon position (U34) of glutamate, lysine and glutamine transfer RNAs is universally modified by thiouridylase into 2-thiouridine (s2U34), which is crucial for precise translation by restricting codon-anticodon wobble during protein synthesis on the ribosome. However, it remains unclear how the enzyme incorporates reactive sulphur into the correct position of the uridine base. Here we present the crystal structures of the MnmA thiouridylase-tRNA complex in three discrete forms, which provide snapshots of the sequential chemical reactions during RNA sulphuration. On enzyme activation, an alpha-helix overhanging the active site is restructured into an idiosyncratic beta-hairpin-containing loop, which packs the flipped-out U34 deeply into the catalytic pocket and triggers the activation of the catalytic cysteine residues. The adenylated RNA intermediate is trapped. Thus, the active closed-conformation of the complex ensures accurate sulphur incorporation into the activated uridine carbon by forming a catalytic chamber to prevent solvent from accessing the catalytic site. The structures of the complex with glutamate tRNA further reveal how MnmA specifically recognizes its three different tRNA substrates. These findings provide the structural basis for a general mechanism whereby an enzyme incorporates a reactive atom at a precise position in a biological molecule.
The biogenesis of 2-thiouridine derivatives at the wobble position 34 of bacterial tRNA requires a sulfur-relay system that involves multiple sulfur mediators. Cysteine is the donor of sulphur group. The type of proteins that compose this relay systems depends on the organism or cell considered. As far as E. coli MnmA is concerned, at least 6 proteins compose this complex relay system (IscS, TusA, TusB, TusC, TusD and TusE). U34 thiolation appears not to depend on prior posttransciptional modification of C5 atom of the uridine ring and thus can occur any time during the biogenesis of X5s2U34. A crystal structure of the MnmA-tRNA complex has been solved at 3.1 angstroms of resolution (
Kambampati et al. 2003)..