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The flhDC operon of Salmonella typhimurium is the master control operon required for the expression of the entire flagellar regulon. The flagellar master operon was placed under the tetracycline-inducible promoter PtetA using the T-POP transposon. Cells containing this construct are motile in the presence of tetracycline and non-motile without inducer present. No flagella were visible under the electron microscope when cells were grown without inducer. The class 1, class 2 and class 3 promoters of the flagellar regulon are temporally regulated. After addition of tetracycline, the class 1 flhDC operon was transcribed immediately. Transcription of flgM (which is transcribed from both class 2 and class 3 promoters) began 15 min after induction. At 20 min after induction, the class 2 fliA promoter became active and intracellular FliA protein levels increased; at 30 min after induction, the class 3 fliC promoter was activated. Induction of fliC gene expression coincides with the appearance of FlgM anti-sigma factor in the growth medium. This also coincides with the completion of hook–basal body structures. Rolling cells first appeared 35 min after induction, and excess hook protein (FlgE) was also found in the growth medium at this time. At 45 min after induction, nascent flagellar filaments became visible in electron micrographs and over 40% of the cells exhibited some swimming behaviour. Multiple flagella assemble and grow on individual cells after induction of the master operon. These results confirm that the flagellar regulatory hierarchy of S. typhimurium is temporally regulated after induction. Both FlgM secretion and class 3 gene expression occur upon completion of the hook–basal body structure. [ABSTRACT FROM AUTHOR]

We analysed all major proteins secreted into culture media from Salmonella typhimurium . Proteins in culture supernatants were collected by trichloroacetic acid precipitation, separated in SDS–polyacrylamide gels and analysed by amino acid sequencing. Wild-type strain SJW1103 cells typically gave rise to nine bands in SDS gels: 89, 67, 58, 52, 50, 42, 40, 35 and (sometimes) 28 kDa. A search of the sequences in the available databases revealed that they were either flagellar proteins or virulence factors. Six of them were flagella specific: FlgK or HAP1 (58 kDa), FliC or flagellin (52 kDa), FliD or HAP2 (50 kDa), FlgE or hook protein (42 kDa), FlgL or HAP3 (35 kDa) and FlgD or hook-cap protein (28 kDa). The other four bands were specific for virulence factors: SipA (89 kDa), SipB (67 kDa), SipC (42 kDa) and InvJ (40 kDa). The 42 kDa band was a mixture of FlgE and SipC. We also analysed secreted proteins from more than 30 flagellar mutants, and they were categorized into four groups according to their band patterns: wild type, mot type, polyhook type and master gene type. Virulence factors were constantly secreted at a higher level in all flagellar mutants except a Δmot (motAB deletion) mutant, in which the amounts were greatly reduced. A new morphological pathway of flagellar biogenesis including protein secretion is presented. [ABSTRACT FROM AUTHOR]

Examines the effect of sodium chloride (NaCl) in the culture media on the flagellar filament elongation of Salmonella typhimurium. Mutations in the hook protein flagellar hook-deficient mutants; Number of hook-basal bodies on cells under various NaCl conditions; Cellular flagellin level; Regulation of flagellin expression by competitive interaction of a sigma factor.

The flagellar hook-basal body (HBB) complex of the Gram-positive bacterium Bacillus subtilis was purified and analysed by electron microscopy, gel electrophoresis, and amino acid sequencing of the major component proteins. The purified HBB complex consisted of the inner (M and S) rings, a rod and a hook. There were no outer (P and L) rings that are found in Gram-negative bacteria. The hook was 15 nm in thickness and 70 nm in length, which is thinner and longer than the hook of Salmonella typhimurium. The hook protein had an apparent molecular mass of 29 kDa, and its N-terminal sequence was identical to that of B. subtilis FlgG, which was previously reported as a rod protein. The sequence of the reported FlgG protein of B. subtilis is more closely related to that of FlgE (the hook protein) rather than FlgG (the rod protein) of S. typhimurium, in spite of the difference of the apparent molecular masses between the two hook proteins (29 kDa versus 42 kDa). The hook-basal body contained six major proteins (with apparent moiecutar masses of 82, 59, 35, 32, 29 and 20 kDa) and two minor proteins (23 kDa and 13 kDa), which consistently appeared from preparation to preparation. The N-terminus of each of these proteins was sequenced. Comparison with protein databases revealed the following polypeptide-gene correspondences: 82 kDa, fliF; 59 kDa, flgK; 35 kDa, orfF; 32 kDa, yqhF; 23 kDa, orf3 of the flaA locus; 20 kDa, flgB and flgC; 13 kDa, not determined. The band at 20 kDa was a mixture of FlgB and FlgC, as revealed by two-dimensional gel analysis. Characteristic features of B. subtilis HBB are discussed in comparison with those of S. typhimiurium. [ABSTRACT FROM AUTHOR]

The bacterial flagellum is a motility apparatus in which a long helical filament - the propeller - is driven by a rotary motor embedded in the cell surface. Out of more than 40 genes required for construction of a fully functional flagellum in Salmonella typhimurium, only 18 gene products have been identified in the mature structure. Some other flagellar proteins play logistical roles during construction, which involves the selective export of flagellar components through a central hole in the flagellum. The whole structure is constructed from base to tip by linear assembly; that is, by adding new components on the growing end, resulting in the distal growth of each substructure. Components of the substructures do not necessarily self-assemble, but often demand the help of other proteins. Recent progress in the understanding of flagellar assembly, which has been most extensively studied in S. typhimurium, is reviewed. [ABSTRACT FROM AUTHOR]