Dataset Information

Helicobacter pylori FlhB function: the FlhB C-terminal homologue HP1575 acts as a "spare part" to permit flagellar export when the HP0770 FlhBCC domain is deleted.

ABSTRACT: In Helicobacter pylori 26695, a gene annotated HP1575 encodes a putative protein of unknown function which shows significant similarity to part of the C-terminal domain of the flagellar export protein FlhB. In Salmonella enterica, this part (FlhB(CC)) is proteolytically cleaved from the full-length FlhB, a processing event that is required for flagellar protein export and, thus, motility. The role of FlhB (HP0770) and its C-terminal homologue HP1575 was studied in H. pylori using a range of nonpolar deletion mutants defective in HP1575, HP0770, and the CC domain of HP0770 (HP0770(CC)). Deletion of HP0770 abolished swimming motility, whereas mutants carrying a deletion of either HP1575 or HP0770(CC) retained their ability to swim. An H. pylori strain containing deletions in both HP1575 and HP0770(CC) was nonmotile and did not produce flagella, suggesting that at least one of the two proteins had to be present for flagellar assembly to occur. Indeed, motility was restored when HP1575 was reintroduced into this strain immediately downstream of, but not fused to, the truncated HP0770 gene. Thus, HP1575 can functionally replace HP0770(CC) in this background. Like FlhB in S. enterica, HP0770 appeared to be proteolytically processed at a conserved NPTH processing site. However, mutation of the proline contained within the NPTH site of HP0770 did not affect motility and flagellar assembly, although it clearly interfered with processing when the protein was heterologously produced in Escherichia coli.

SUBMITTER: Wand ME

PROVIDER: S-EPMC1636261 | biostudies-literature | 2006 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Helicobacter pylori FlhB function: the FlhB C-terminal homologue HP1575 acts as a "spare part" to permit flagellar export when the HP0770 FlhBCC domain is deleted.

Wand Matthew E ME Sockett R Elizabeth RE Evans Katy J KJ Doherty Neil N Sharp Paul M PM Hardie Kim R KR Winzer Klaus K

Journal of bacteriology 20061101 21

In Helicobacter pylori 26695, a gene annotated HP1575 encodes a putative protein of unknown function which shows significant similarity to part of the C-terminal domain of the flagellar export protein FlhB. In Salmonella enterica, this part (FlhB(CC)) is proteolytically cleaved from the full-length FlhB, a processing event that is required for flagellar protein export and, thus, motility. The role of FlhB (HP0770) and its C-terminal homologue HP1575 was studied in H. pylori using a range of nonp ...[more]

PMID: 17050924

Similar Datasets

Project description:The bacterial type III export apparatus is found in the flagellum and in the needle complex of some pathogenic Gram-negative bacteria. In the needle complex its function is to secrete effector proteins for infection into Eukaryotic cells. In the bacterial flagellum it exports specific proteins for the building of the flagellum during its assembly. The export apparatus is composed of about five membrane proteins and three soluble proteins. The mechanism of the export apparatus is not fully understood. The five membrane proteins are well conserved and essential. Here a cross-complementation assay was performed: substituting in the flagellar system of Salmonella one of these membrane proteins, FlhB, by the FlhB ortholog from Aquifex aeolicus (an evolutionary distant hyperthermophilic bacteria) or a chimeric protein (AquSalFlhB) made by the combination of the trans-membrane domain of A. aeolicus FlhB with the cytoplasmic domain of Salmonella FlhB dramatically reduced numbers of flagella and motility. From cells expressing the chimeric AquSalFlhB protein, suppressor mutants with enhanced motility were isolated and the mutations were identified using whole genome sequencing. Gain-of-function mutations were found in the gene encoding FlhA, another membrane protein of the type III export apparatus. Also, mutations were identified in genes encoding 4-hydroxybenzoate octaprenyltransferase, ubiquinone/menaquinone biosynthesis methyltransferase, and 4-hydroxy-3-methylbut-2-en-1-yl diphosphate synthase, which are required for ubiquinone biosynthesis. The mutations were shown by reversed-phase high performance liquid chromatography to reduce the quinone pool of the cytoplasmic membrane. Ubiquinone biosynthesis could be restored for the strain bearing a mutated gene for 4-hydroxybenzoate octaprenyltransferase by the addition of excess exogenous 4-hydroxybenzoate. Restoring the level of ubiquinone reduced flagella biogenesis with the AquSalFlhB chimera demonstrating that the respiratory chain quinone pool is responsible for this phenomenon.

Project description:FlhA and FlhB are transmembrane proteins of the flagellar type III protein export apparatus, and their C-terminal cytoplasmic domains (FlhAC and FlhBC) coordinate flagellar protein export with assembly. FlhBC undergoes autocleavage between Asn-269 and Pro-270 in a well-conserved NPTH loop located between FlhBCN and FlhBCC polypeptides and interacts with the C-terminal domain of the FliK ruler when the length of the hook has reached about 55 nm in Salmonella As a result, the flagellar protein export apparatus switches its substrate specificity, thereby terminating hook assembly and initiating filament assembly. The mechanism of export switching remains unclear. Here, we report the role of FlhBC cleavage in the switching mechanism. Photo-cross-linking experiments revealed that the flhB(N269A) and flhB(P270A) mutations did not affect the binding affinity of FlhBC for FliK. Genetic analysis of the flhB(P270A) mutant revealed that the P270A mutation affects a FliK-dependent conformational change of FlhBC, thereby inhibiting the substrate specificity switching. The flhA(A489E) mutation in FlhAC suppressed the flhB(P270A) mutation, suggesting that an interaction between FlhBC and FlhAC is critical for the export switching. We propose that the interaction between FliKC and a cleaved form of FlhBC promotes a conformational change in FlhBC responsible for the termination of hook-type protein export and a structural remodeling of the FlhAC ring responsible for the initiation of filament-type protein export.IMPORTANCE The flagellar type III protein export apparatus coordinates protein export with assembly, which allows the flagellum to be efficiently built at the cell surface. Hook completion is an important morphological checkpoint for the sequential flagellar assembly process. The protein export apparatus switches its substrate specificity from the hook protein to the filament protein upon hook completion. FliK, FlhB, and FlhA are involved in the export-switching process, but the mechanism remains a mystery. By analyzing a slow-cleaving flhB(P270A) mutant, we provide evidence that an interaction between FliK and FlhB induces conformational rearrangements in FlhB, followed by a structural remodeling of the FlhA ring structure that terminates hook assembly and initiates filament formation.

Dataset Information

Helicobacter pylori FlhB function: the FlhB C-terminal homologue HP1575 acts as a "spare part" to permit flagellar export when the HP0770 FlhBCC domain is deleted.

Publications

Helicobacter pylori FlhB function: the FlhB C-terminal homologue HP1575 acts as a "spare part" to permit flagellar export when the HP0770 FlhBCC domain is deleted.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets