The FliP and FliR proteins of Salmonella typhimurium, putative components of the type III flagellar export apparatus, are located in the flagellar basal body.
Most of the structural components of the flagellum of Salmonella typhimurium are exported through a flagellum-specific pathway, which is a member of the family of type III secretory pathways. The export apparatus for this process is poorly understood. A previous study has shown that two proteins, about 23 and 26 kDa in size and of unknown genetic origin, are incorporated into the flagellar basal body at a very early stage of flagellar assembly. In the present study, we demonstrate that these basal body proteins are FliP (in its mature form after signal peptide cleavage) and FliR respectively. Both of these proteins have homologues in other type III secretion systems. By placing a FLAG epitope tag on FliR and the MS-ring protein FliF and immunoblotting isolated hook basal body complexes with anti-FLAG monoclonal antibody, we estimate (using the FLAG-tagged FliF as an internal reference) that the stoichiometry of FliR is fewer than three copies per basal body. An independent estimate of stoichiometry was made using data from an earlier quantitative radiolabelling analysis, yielding values of around four or five subunits per basal body for FliP and around one subunit per basal body for FliR. Immunoelectron microscopy using anti-FLAG antibody and gold-protein A suggests that FliR is located near the MS ring. We propose that the flagellar export apparatus contains FliP and FliR and that this apparatus is embedded in a patch of membrane in the central pore of the MS ring.
Fan
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F
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520-8114, USA.
Ohnishi
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K
Francis
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NR
Macnab
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RM
D001345:Autoradiography; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000939:Epitopes / Q000235:genetics / Q000378:metabolism; D005407:Flagella / Q000378:metabolism*; D006639:Histidine / Q000378:metabolism; D015151:Immunoblotting; D008565:Membrane Proteins; D016253:Microscopy, Immunoelectron; D009842:Oligopeptides; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D012486:Salmonella typhimurium / Q000378:metabolism*
D001426:Bacterial Proteins; D000939:Epitopes; C085248:FliR protein, Bacteria; D008565:Membrane Proteins; D009842:Oligopeptides; D010455:Peptides; C074890:FliP protein, Bacillus; D006639:Histidine; C085706:FLAG peptide