PMID:16246842
| Citation |
Ferris, HU, Furukawa, Y, Minamino, T, Kroetz, MB, Kihara, M, Namba, K and Macnab, RM (2005) FlhB regulates ordered export of flagellar components via autocleavage mechanism. J. Biol. Chem. 280:41236-42 |
|---|---|
| Abstract |
The bacterial flagellum is a predominantly cell-external super-macromolecular construction whose structural components are exported by a flagellum-specific export apparatus. One of the export apparatus proteins, FlhB, regulates the substrate specificity of the entire apparatus; i.e. it has a role in the ordered export of the two main groups of flagellar structural proteins such that the cell-proximal components (rod-/hook-type proteins) are exported before the cell-distal components (filament-type proteins). The controlled switch between these two export states is believed to be mediated by conformational changes in the structure of the C-terminal cytoplasmic domain of FlhB (FlhB(C)), which is consistently and specifically cleaved into two subdomains (FlhB(CN) and FlhB(CC)) that remain tightly associated with each other. The cleavage event has been shown to be physiologically significant for the switch. In this study, the mechanism of FlhB cleavage has been more directly analyzed. We demonstrate that cleavage occurs in a heterologous host, Saccharomyces cerevisiae, deficient in vacuolar proteinases A and B. In addition, we find that cleavage of a slow-cleaving variant, FlhB(C)(P270A), is stimulated in vitro at alkaline pH. We also show by analytical gel-filtration chromatography and analytical ultracentrifugation experiments that both FlhB(C) and FlhB(C)(P270A) are monomeric in solution, and therefore self-proteolysis is unlikely. Finally, we provide evidence via peptide analysis and FlhB cleavage variants that the tertiary structure of FlhB plays a significant role in cleavage. Based on these results, we propose that FlhB cleavage is an autocatalytic process. |
| Links |
PubMed Online version:10.1074/jbc.M509438200 |
| Keywords |
Bacterial Proteins/chemistry; Bacterial Proteins/physiology; Catalysis; Chromatography, Gel; Electrophoresis, Polyacrylamide Gel; Escherichia coli/metabolism; Flagella/metabolism; Fungal Proteins/chemistry; Gene Expression Regulation, Bacterial; Gene Expression Regulation, Fungal; Hydrogen-Ion Concentration; Immunoblotting; Membrane Proteins/physiology; Models, Biological; Models, Chemical; Mutation; Peptides/chemistry; Plasmids/metabolism; Protein Binding; Protein Conformation; Protein Structure, Tertiary; Saccharomyces cerevisiae/metabolism; Salmonella/metabolism; Substrate Specificity; Succinimides/chemistry; Ultracentrifugation |
| edit table |
Significance
You can help EcoliWiki by summarizing why this paper is useful
Useful Materials and Methods
You can help Ecoliwiki by describing the useful materials (strains, plasmids, antibodies, etc) described in this paper.
Annotations
<annotationlinks/>
EcoliWiki Links
Add links to pages that link here (e.g. gene, product, method pages)
References
See Help:References for how to manage references in EcoliWiki.