Purification and characterization of the flagellar hook–basal body complex of <i>Bacillus subtilis</i>

Kubori, Tomoko; Okumura, Mitsumasa; Kobayashi, N.; Nakamura, Dai; Iwakura, Masahiro; Aizawa, Shin‐Ichi

doi:10.1046/j.1365-2958.1997.3341714.x

Cited by 37 publications

(39 citation statements)

References 29 publications

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“…In support of this idea, several studies focused on the functional properties of flagellum-and contact-dependent TTSSs have revealed common themes leading to the proposal of a unifying model for how substrates are recognized and transported (24). Further support for a unifying model comes from the direct examination of flagellum-and contact-dependent TTSS structures, which has revealed striking images of the protein targeting apparatus with highly conserved architectures (1,5,27,28,51). The assembly of flagellum-and contact-dependent TTSS structures appears to occur by a similar process (50).…”

Section: Discussionmentioning

confidence: 87%

“…It has been hypothesized that yplB may encode an accessory protein or chaperone required for YplA secretion, but this function has not been established. Located immediately upstream of yplA are Ϫ35 and Ϫ10 promoter regions that share identity with the consensus sequence of known 28 -dependent promoters and expression of yplA was shown to require the function of fliA (fliA encodes 28 ) (45,46). This indicates that yplA belongs to the flagellar regulon and that YplA is not made until the hook-basal body is complete.…”

mentioning

confidence: 99%

“…During the final stages of flagellar biogenesis the hook-basal body serves as both the machinery for the export of filament subunits and the base for filament assembly (30). Coordination of flagellar genes that encode components of the filament, the rotary motor, and the chemosensory system are expressed from 28 -dependent promoters that provide a regulatory mechanism that ensures these components are not synthesized until after the initial flagellar structure is complete (25).…”

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confidence: 99%

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YplA Is Exported by the Ysc, Ysa, and Flagellar Type III Secretion Systems ofYersinia enterocolitica

2002

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Yersinia enterocolitica maintains three different pathways for type III protein secretion. Each pathway requires the activity of a specific multicomponent apparatus or type III secretion system (TTSS). Two of the TTSSs are categorized as contact-dependent systems which have been shown in a number of different symbiotic and pathogenic bacteria to influence interactions with host organisms by targeting effector proteins into the cytosol of eukaryotic cells. The third TTSS is required for the assembly of flagella and the secretion of the phospholipase YplA, which has been implicated in Y. enterocolitica virulence. In this study, YplA was expressed from a constitutive promoter in strains that contained only a single TTSS. It was determined that each of the three TTSSs is individually sufficient for YplA secretion. Environmental factors such as temperature, calcium availability, and sodium chloride concentration affected the contribution of each system to extracellular protein secretion and, under some conditions, more than one TTSS appeared to operate simultaneously. This suggests that some proteins might normally be exported by more than one TTSS in Y. enterocolitca.

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Section: Discussionmentioning

confidence: 87%

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confidence: 99%

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confidence: 99%

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YplA Is Exported by the Ysc, Ysa, and Flagellar Type III Secretion Systems ofYersinia enterocolitica

2002

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“…For example, Bacillus subtilis, one of the best-characterized Gram-positive bacteria belonging to Firmicutes, encodes neither dual-nor single-domain FlgJ homolog (Kunst et al, 1997), though it produces flagella actively (Kubori et al, 1997a). This implies that the rod assembly machinery in these bacterial species may be substantially different from those in proteobacteria.…”

Section: Discussionmentioning

confidence: 95%

Plasticity of the domain structure in FlgJ, a bacterial protein involved in flagellar rod formation

2006

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Bacterial flagellar rod structure is built across the peptidoglycan (PG) layer. A Salmonella enterica flagellar protein FlgJ is believed to consist of two functional domains, the N-terminal half acting as a scaffold or cap essential for rod assembly and the C-terminal half acting as a PG hydrolase (PGase) that makes a hole in the PG layer to facilitate rod penetration. In this study, molecular data analyses were conducted on FlgJ data sets sampled from a variety of bacterial species, and three types of FlgJ homologs were identified: (i) "canonical dual-domain" type found in β-and γ-proteobacteria that has a domain for one of the PGases, acetylmuramidase (Acm), at the C terminus, (ii) "non-canonical dual-domain" type found in the genus Desulfovibrio (δ-proteobacteria) that bears a domain for another PGase, M23/M37-family peptidase (Pep), at the C terminus and (iii) "singledomain" type found in phylogenetically diverged lineages that lacks the Acm or Pep domain. FlgJ phylogeny, together with the domain architecture, suggested that the single-domain type was the original form of FlgJ and the canonical dualdomain type had evolved from the single-domain type by fusion of the Acm domain to its C terminus in the common ancestor of β-and γ-proteobacteria. The noncanonical dual-domain type may have been formed by fusion of the Pep domain to the single-domain type in the ancestor of Desulfovibrio. In some lineages of γ-proteobacteria, the Acm domain appeared to be lost secondarily from the dual-domain type FlgJ to yield again a single-domain type one. To rationalize the underlying mechanism that gave rise to the two different types of dual-domain FlgJ homologs, we propose a model assuming the lineage-specific co-option of flagellum-specific PGase from diverged housekeeping PGases in bacteria.

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“…2B). Coomassie blue staining confirmed that outer membrane proteins located toward the bottom of the gradient, and immunoblotting revealed both the N⌬120-140 and N⌬130-140 in the inner-membrane fractions colocalizing with the NADH oxidase and flagellar C-ring component FliM (27,28).…”

Section: Dominant-negative Export-defective Chaperone Variants Localimentioning

confidence: 95%

Docking of cytosolic chaperone-substrate complexes at the membrane ATPase during flagellar type III protein export

Thomas

Stafford

Hughes

2004

Proc. Natl. Acad. Sci. U.S.A.

130

155

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Bacterial type III protein export underlies flagellum assembly and delivery of virulence factors into eukaryotic cells. The sequence of protein interactions underlying the export pathway are poorly characterized; in particular, it is not known how chaperoned substrates in the cytosol are engaged by the membrane-localized export apparatus. We have identified a stalled intermediate export complex in the flagellar type III export pathway of Salmonella typhimurium by generating dominant-negative chaperone variants that are export-defective and arrest flagellar assembly in the wild-type bacterium. These chaperone variants bound their specific export substrates strongly and severely reduced their export. They also attenuated export of other flagellar proteins, indicating that inhibition occurs at a common step in the pathway. Unlike the cytosolic wild-type chaperone, the variants localized to the inner membrane, but not in the absence of the flagellar type III export apparatus. Membrane localization persisted in fliOPQR, flhB, flhA, fliJ, and fliH null mutants lacking specific flagellar export components but depended on the presence of the membrane-associated ATPase FliI. After expression of the variant chaperones in Salmonella, a stalled intermediate export complex, which contained chaperone, substrate, and the FliI ATPase with its regulator FliH, was isolated. Neither chaperone nor substrate alone was able to interact with liposome-associated FliI, but the chaperone-substrate-FliI(FliH) complex was assembled when chaperone was prebound to its substrate. Our data establish a key event in the type III protein export mechanism, docking of the cytosolic chaperone-substrate complex at the ATPase of the membrane-export apparatus.

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Purification and characterization of the flagellar hook–basal body complex of Bacillus subtilis

Cited by 37 publications

References 29 publications

YplA Is Exported by the Ysc, Ysa, and Flagellar Type III Secretion Systems ofYersinia enterocolitica

YplA Is Exported by the Ysc, Ysa, and Flagellar Type III Secretion Systems ofYersinia enterocolitica

Plasticity of the domain structure in FlgJ, a bacterial protein involved in flagellar rod formation

Docking of cytosolic chaperone-substrate complexes at the membrane ATPase during flagellar type III protein export

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