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Bacteria swim in liquid environments by means of a complex rotating structure known as the flagellum. Approximately 40 proteins are required for the assembly and functionality of this structure. Rhodobacter sphaeroides has two flagellar systems. One of these systems has been shown to be functional and is required for the synthesis of the well-characterized single subpolar flagellum, while the other was found only after the genome sequence of this bacterium was completed. In this work we found that the second flagellar system of R. sphaeroides can be expressed and produces a functional flagellum. In many bacteria with two flagellar systems, one is required for swimming, while the other allows movement in denser environments by producing a large number of flagella over the entire cell surface. In contrast, the second flagellar system of R. sphaeroides produces polar flagella that are required for swimming. Expression of the second set of flagellar genes seems to be positively regulated under anaerobic growth conditions. Phylogenic analysis suggests that the flagellar system that was initially characterized was in fact acquired by horizontal transfer from a ␥-proteobacterium, while the second flagellar system contains the native genes. Interestingly, other ␣-proteobacteria closely related to R. sphaeroides have also acquired a set of flagellar genes similar to the set found in R. sphaeroides, suggesting that a common ancestor received this gene cluster.The bacterial flagellum is a complex protein structure which consists of a long helical filament that is connected through a flexible linker known as the hook to an H ϩ -or Na ϩ -driven rotary motor (29). Rotation of the filament produces thrust that allows the cell to swim in liquid or semisolid medium (6). Bacteria perform taxis by controlling the frequency of reorientation, which is commonly regulated by a two-component signal transduction system that senses an environmental stimulus (2, 3, 7). Several reorientation mechanisms have been described for different bacteria (35). In Escherichia coli and Salmonella enterica serovar Typhimurium, in which the flagellar system has been studied more extensively, more than 40 proteins are required for flagellar synthesis and functioning (29). Expression of the flagellar genes is regulated in a hierarchical pattern which results from coordination of flagellar gene expression at the transcriptional or posttranscriptional level with one or more structural checkpoints in flagellum biogenesis (31). This tight regulation has probably evolved to avoid unnecessary synthesis of the large number of flagellar protein subunits required for this structure. The high energetic cost required for the synthesis and functioning of the flagellum is compensated for by the selective advantage conferred by motility. Accordingly, motility seems to be important in several processes, such as colonization, pathogenesis, dispersion, and competition for resources (37). When the growth medium is too dense to allow swimming, many bacteria differentiate in...

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FliI is a Salmonella typhimurium protein that is needed for flagellar assembly and may be involved in a specialized protein export pathway that proceeds without signal peptide cleavage. FliT shows extensive sequence similarity to the catalytic 13 subunit of the FoF1 ATPase (A. P. Vogler, M. Homma, V. M. Irikura, and R. M. Macnab, J. Bacteriol. 173:3564-3572, 1991). It is even more similar to the Spa47 protein of Shigella flexneri (M. M. Venkatesan, J. M. Buysse, and E. V. Oaks, J. Bacteriol. 174:1990-2001) and the HrpB6 protein of Xanthomonas campestris (S. Fenselau, I. Balbo, and U. Bonas, Mol. Plant-Microbe Interact. 5:390-396, 1992), which are believed to play a role in the export of virulence proteins. Site-directed mutagenesis of residues in FliI that correspond to catalytically important residues in the F1 ,3 subunit resulted in loss of flagellation, supporting the hypothesis that Fliu is an ATPase. FliT was overproduced and purified almost to homogeneity. It demonstrated ATP binding but not hydrolysis. An antibody raised against FliI permitted detection of the protein in wild-type cells and an estimate of about 1,500 subunits per cell. An antibody directed against the F1 P subunit ofEscherichia coli cross-reacted with Flil, confirming that the proteins are structurally related. The relationship between three proteins involved in flagellar assembly (FliI, FlhA, and FliP) and homologs in a variety of virulence systems is discussed.The flagellum of Salmonella typhimurium and those of many other bacteria consist of at least three distinct structures: the filament and hook (both of which are completely external to the cell) and the basal body (which spans the cytoplasmic membrane, the periplasmic space, and the outer membrane) (see reference 36 for a review). Thus, many flagellar proteins, following their synthesis inside the cell, need to be exported from the cytoplasm. Only two of these (FlgH and FlgI, from which the outer pair of basal-body rings are constructed) appear to be exported by the conventional signal peptide-dependent pathway (22,26). The rest do not undergo cleavage of a signal peptide and are presumed to be exported by a unique flagellum-specific pathway (36). The major protein in this class, flagellin, is known to assemble at the distal end of the growing filament (10, 25), and it is presumed that the proteins constituting the other substructures such as the rod and hook are also assembled by distal addition, since they are related to flagellin (20, 23) and the order of assembly of these substructures is from cell proximal to cell distal (first rod, then hook, and finally filament) (27,31,48,49). The physical path for travel of subunits to their destination is almost certainly through the hollow channel that exists in the structure (38,42 vicinity of the flagellar base, that transports the proteins across the plane of the cell membrane (although not through the bulk membrane itself) into this channel. This export apparatus must be selective in the proteins it recognizes, but a flagellum-speci...