title: A response regulatory protein with the site of phosphorylation blocked by an arginine interaction: crystal structure of Spo0F from Bacillus subtilis.
abstract: Spo0F is a secondary messenger in the "two-component" system controlling the sporulation of Bacillus subtilis. Spo0F, like the chemotaxis protein CheY, is a single-domain protein homologous to the N-terminal activator domain of the response regulators. We recently reported the crystal structure of a phosphatase-resistant mutant Y13S of Spo0F with Ca2+ bound in the active site. The crystal structure of wild-type Spo0F in the absence of a metal ion is presented here. A comparison of the two structures reveals that the cation induces significant changes in the active site. In the present wild-type structure, the carboxylate of Asp11 points away from the center of the active site, whereas when coordinated to the Ca2+, as in the earlier structure, it points toward the active site. In addition, Asp54, the site of phosphorylation, is blocked by a salt bridge interaction of an Arg side chain from a neighboring molecule. From fluorescence quenching studies with Spo0F Y13W, we found that only the amino acid Arg binds to Spo0F in a saturable manner (Kd = 15 mM). This observation suggests that a small molecule with a shape complementary to the active site and having a guanidinium group might inhibit phosphotransfer between response regulators and their cognate histidine kinases.
authors.affiliation: Department of Biology, University of California at San Diego, La Jolla, California 92093-0359, USA.
authors.forename: M
authors.initials: M
authors.lastname: Madhusudan
authors.forename: J
authors.initials: J
authors.lastname: Zapf
authors.forename: J A
authors.initials: JA
authors.lastname: Hoch
authors.forename: J M
authors.initials: JM
authors.lastname: Whiteley
authors.forename: N H
authors.initials: NH
authors.lastname: Xuong
authors.forename: K I
authors.initials: KI
authors.lastname: Varughese
mesh_terms: D019943:Amino Acid Substitution; D001120:Arginine; D001412:Bacillus subtilis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D018360:Crystallography, X-Ray; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015290:Second Messenger Systems; D055672:Static Electricity
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; C080951:Spo0F protein, Bacillus subtilis; D001120:Arginine; D002118:Calcium