Domain Organization and Function of Salmonella FliK, a Flagellar Hook-length Control Protein
| Autor: | Tohru Minamino, Yumiko Saijo-Hamano, Yukio Furukawa, Bertha González-Pedrajo, Robert M. Macnab, Keiichi Namba |
|---|---|
| Rok vydání: | 2004 |
| Předmět: |
Cytoplasm
Conformational change Proteolysis Molecular Sequence Data Flagellum Mass Spectrometry Substrate Specificity Flik Bacterial Proteins Salmonella Structural Biology medicine Trypsin Amino Acid Sequence Molecular Biology Polyacrylamide gel electrophoresis Sequence Homology Amino Acid medicine.diagnostic_test Chemistry Escherichia coli Proteins Flagellar hook Biological Transport Protein Structure Tertiary Cell biology Biochemistry Flagella Chromatography Gel Ultracentrifugation Function (biology) |
| Zdroj: | Journal of Molecular Biology. 341:491-502 |
| ISSN: | 0022-2836 |
| DOI: | 10.1016/j.jmb.2004.06.012 |
| Popis: | Salmonella hook-length control protein FliK, which consists of 405 amino acid residues, switches substrate specificity of the type III flagellar protein export apparatus from rod/ hook-type to filament-type by causing a conformational change in the cytoplasmic domain of FlhB (FlhB C ) upon completion of the hook assembly. An N-terminal region of FliK contains an export signal, and a highly conserved C-terminal region consisting of amino acid residues 265–405 (FliK (265–405) ) is directly involved in the switching of FlhB C . Here, we have investigated the structural properties of FliK. Gel filtration chromatography, multi-angle light scattering and analytical ultracentrifugation showed that FliK is monomeric in solution and has an elongated shape. Limited proteolysis showed that FliK consists of two domains, the N-terminal (FliK N ) and C-terminal domains (FliK C ), and that the first 203 and the last 35 amino acid residues are partially unfolded and subjected to proteolysis. Both FliK N and FliK C are more globular than full-length FliK, suggesting that these domains are connected in tandem. Overproduced His-FliK (199–405) failed to switch export specificity of the export apparatus. Affinity blotting revealed that FlhB C binds to FliK and FliK (1–147) , but not to FliK (265–405) . Based on these results, we propose that FliK N within the central channel of the hook-basal body during the export of FliK is the sensor and transmitter of hook completion information and that the binding interaction of FliK C to FlhB C is structurally regulated by FliK N so as to occur only when the hook has reached a preset length. The conformational flexibility of FliK C may play a role in interfering with switching at an inappropriate point of flagellar assembly. |
| Databáze: | OpenAIRE |
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