3HFY

Mutant of tRNA-guanine transglycosylase (K52M)

Summary for 3HFY

• Entry DOI: 10.2210/pdb3hfy/pdb
• Related: 1P0D 1P0E 3EOS
• Descriptor: Queuine tRNA-ribosyltransferase, ZINC ION (3 entities in total)
• Functional Keywords: tgt, dimer interface, mutation, glycosyltransferase, metal-binding, queuosine biosynthesis, transferase, trna processing, zinc
• Biological source: Zymomonas mobilis
• Total number of polymer chains: 1
• Total formula weight: 42993.13

Authors

Ritschel, T.,Klebe, G. (deposition date: 2009-05-13, release date: 2009-08-18, Last modification date: 2024-05-29)

Primary citation

Ritschel, T.,Atmanene, C.,Reuter, K.,Van Dorsselaer, A.,Sanglier-Cianferani, S.,Klebe, G.
An Integrative Approach Combining Noncovalent Mass Spectrometry, Enzyme Kinetics and X-ray Crystallography to Decipher Tgt Protein-Protein and Protein-RNA Interaction
J.Mol.Biol., 393:833-847, 2009

PubMed Abstract: The tRNA-modifying enzyme tRNA-guanine transglycosylase (Tgt) is a putative target for new selective antibiotics against Shigella bacteria. The formation of a Tgt homodimer was suggested on the basis of several crystal structures of Tgt in complex with RNA. In the present study, noncovalent mass spectrometry was used (i) to confirm the dimeric oligomerization state of Tgt in solution and (ii) to evidence the binding stoichiometry of the complex formed between Tgt and its full-length substrate tRNA. To further investigate the importance of Tgt protein-protein interaction, point mutations were introduced into the dimer interface in order to study their influence on the formation of the catalytically active complex. Enzyme kinetics revealed a reduced catalytic activity of these mutated variants, which could be related to a destabilization of the dimer formation as evidenced by both noncovalent mass spectrometry and X-ray crystallography. Finally, the effect of inhibitor binding was investigated by noncovalent mass spectrometry, thus providing the binding stoichiometries of Tgt:inhibitor complexes and showing competitive interactions in the presence of tRNA. Inhibitors that display an influence on the formation of the dimer interface in the crystal structure are promising candidates to alter the protein-protein interaction, which could provide a new way to inhibit Tgt.

PubMed: 19627989
DOI: 10.1016/j.jmb.2009.07.040

Experimental method

X-RAY DIFFRACTION (2 Å)