The importance of charge in perturbing the aromatic glue stabilizing the protein-protein interface of homodimeric tRNA-guanine transglycosylase

Author:

Nguyen AndreasORCID,Nguyen DzungORCID,Nguyen Tran Xuan Phong,Sebastiani Maurice,Dörr Stefanie,Hernandez-Alba OscarORCID,Debaene FrançoisORCID,Cianférani SarahORCID,Heine AndreasORCID,Klebe GerhardORCID,Reuter KlausORCID

Abstract

AbstractBacterial tRNA-guanine transglycosylase (Tgt) is involved in the biosynthesis of the modified tRNA nucleoside queuosine present in the anticodon wobble position of tRNAs specific for aspartate, asparagine, histidine and tyrosine. Inactivation of thetgtgene leads to decreased pathogenicity ofShigellabacteria. Therefore, Tgt constitutes a putative target for Shigellosis drug therapy. Since only active as homodimer, interference with dimer-interface formation may, in addition to active-site inhibition, provide further means to disable this protein. A cluster of four aromatic residues seems important to stabilize the homodimer. We mutated residues of this aromatic cluster and analyzed each exchange with respect to dimer and thermal stability or enzyme activity applying native mass spectrometry, thermal shift assay, enzyme kinetics, and X-ray crystallography. Our structural studies indicate strong influence of pH on homodimer stability. Obviously, protonation of a histidine within the aromatic cluster promotes the collapse of an essential structural motif within the dimer interface at slightly acidic pH.TOC GraphicFor table of contents use only.

Publisher

Cold Spring Harbor Laboratory

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