title: In vivo sulfhydryl modification of the ligand-binding site of Tsr, the Escherichia coli serine chemoreceptor.
abstract: The Escherichia coli chemoreceptor Tsr mediates an attractant response to serine. We substituted Cys for Thr-156, one of the residues involved in serine sensing. The mutant receptor Tsr-T156C retained serine- and repellent-sensing abilities. However, it lost serine-sensing ability when it was treated in vivo with sulfhydryl-modifying reagents such as N-ethylmaleimide (NEM). Serine protected Tsr-T156C from these reagents. We showed that [3H]NEM bound to Tsr-T156C and that binding decreased in the presence of serine. By pretreating cells with serine and cold NEM, Tsr-T156C was selectively labeled with radioactive NEM. These results are consistent with the location of Thr-156 in the serine-binding site. Chemical modification of the Tsr ligand-binding site provides a basis for simple purification and should assist further in vivo and in vitro investigations of this chemoreceptor protein.
authors.affiliation: Department of Molecular Biology, Faculty of Science, Nagoya University, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Iwama
authors.forename: I
authors.initials: I
authors.lastname: Kawagishi
authors.forename: S
authors.initials: S
authors.lastname: Gomi
authors.forename: M
authors.initials: M
authors.lastname: Homma
authors.forename: Y
authors.initials: Y
authors.lastname: Imae
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003545:Cysteine / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D012694:Serine / Q000378:metabolism; D013912:Threonine / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D008024:Ligands; D008565:Membrane Proteins; C057635:Tsr protein, Bacteria; D013912:Threonine; D012694:Serine; D003545:Cysteine; D005033:Ethylmaleimide