Protein-protein interaction between Bacillus stearothermophilus tyrosyl-tRNA synthetase subdomains revealed by a bacterial two-hybrid system.
We have recently developed a bacterial two-hybrid system (BACTH), based on functional complementation between two fragments of the catalytic domain of Bordetella pertussis adenylate cyclase (AC), that allows an easy in vivo screening and selection of functional interactions between two proteins in Escherichia coli. In this work, we have further explored the potentialities of the BACTH system to study protein-protein interactions, using as a model, the interactions between various subdomains of the dimeric tyrosyl-tRNA synthetase (TyrRS) of Bacillus stearothermophilus. Using the BACTH system we confirmed the known interactions of the alpha/beta domains and those between the alpha/beta domain and the alpha domain that could be anticipated from the three-dimensional structure of TyrRS. Interestingly, the BACTH system revealed the unexpected interaction between the TyrRS alpha domains which is presumably mediated by a pseudo-leucine zipper motif. This study illustrates the interest of the bacterial two-hybrid system to delineate interacting domains of proteins and shows that it can reveal interactions that occur in vivo and that were not anticipated from the three-dimensional structure of the protein of interest.
Karimova
G
G
Unité de Biochimie Cellulaire, CNRS URA 2185, Institut Pasteur, Paris, France. karimova@pasteur.fr
Ullmann
A
A
Ladant
D
D
D000262:Adenylyl Cyclases / Q000235:genetics; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001483:Base Sequence; D001886:Bordetella pertussis / Q000201:enzymology; D019281:Dimerization; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017434:Protein Structure, Tertiary; D020798:Two-Hybrid System Techniques; D014443:Tyrosine / Q000378:metabolism; D014448:Tyrosine-tRNA Ligase / Q000235:genetics / Q000378:metabolism*
D014443:Tyrosine; D000262:Adenylyl Cyclases; D014448:Tyrosine-tRNA Ligase