Topology of the VirB4 C Terminus in the Agrobacterium tumefaciens VirB/D4 Type IV Secretion System

Draper, Olga; Middleton, Rebecca; Doucleff, Michaeleen; Zambryski, Patricia

doi:10.1074/jbc.m606403200

Cited by 23 publications

(28 citation statements)

References 42 publications

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“…TrwK oligomerization will promote pilus assembly and stabilization of the core components, TrwG and TrwE (VirB8 and VirB10, respectively), in an ATPase-independent process (45). Once anchored to the membrane, VirB4 (TrwK) will interact with VirB11 (TrwD) (13,45). TrwD (VirB11) is a hexameric ATPase that could be involved in the unfolding of the proteins or toxins to be transported (33,44), whereas TrwK will energize substrate transport (45).…”

Section: Discussionmentioning

confidence: 99%

ATPase Activity and Oligomeric State of TrwK, the VirB4 Homologue of the Plasmid R388 Type IV Secretion System

et al. 2008

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Type IV secretion systems (T4SS) mediate the transfer of DNA and protein substrates to target cells. TrwK, encoded by the conjugative plasmid R388, is a member of the VirB4 family, comprising the largest and most conserved proteins of T4SS. VirB4 was suggested to be an ATPase involved in energizing pilus assembly and substrate transport. However, conflicting experimental evidence concerning VirB4 ATP hydrolase activity was reported. Here, we demonstrate that TrwK is able to hydrolyze ATP in vitro in the absence of its potential macromolecular substrates and other T4SS components. The kinetic parameters of its ATPase activity have been characterized. The TrwK oligomerization state was investigated by analytical ultracentrifugation and electron microscopy, and its effects on ATPase activity were analyzed. The results suggest that the hexameric form of TrwK is the catalytically active state, much like the structurally related protein TrwB, the conjugative coupling protein.

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Section: Discussionmentioning

confidence: 99%

ATPase Activity and Oligomeric State of TrwK, the VirB4 Homologue of the Plasmid R388 Type IV Secretion System

et al. 2008

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“…Since LTs are ubiquitous in most peptidoglycan-containing eubacteria, they provide a potential target for new antibacterial drugs. LTs can be associated physically with components of protein secretion systems, as shown for VirB1 of the type IV secretion systems of Agrobacterium tumefaciens and Brucella suis (145,586). This might ensure that peptidoglycan is degraded only locally.…”

Section: Contribution Of Peptidoglycan-degrading Enzymesmentioning

confidence: 99%

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

Büttner

2012

Microbiol Mol Biol Rev

366

482

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SUMMARY Flagellar and translocation-associated type III secretion (T3S) systems are present in most Gram-negative plant- and animal-pathogenic bacteria and are often essential for bacterial motility or pathogenicity. The architectures of the complex membrane-spanning secretion apparatuses of both systems are similar, but they are associated with different extracellular appendages, including the flagellar hook and filament or the needle/pilus structures of translocation-associated T3S systems. The needle/pilus is connected to a bacterial translocon that is inserted into the host plasma membrane and mediates the transkingdom transport of bacterial effector proteins into eukaryotic cells. During the last 3 to 5 years, significant progress has been made in the characterization of membrane-associated core components and extracellular structures of T3S systems. Furthermore, transcriptional and posttranscriptional regulators that control T3S gene expression and substrate specificity have been described. Given the architecture of the T3S system, it is assumed that extracellular components of the secretion apparatus are secreted prior to effector proteins, suggesting that there is a hierarchy in T3S. The aim of this review is to summarize our current knowledge of T3S system components and associated control proteins from both plant- and animal-pathogenic bacteria.

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“…TrwB was extensively characterized as a DNA-dependent ATPase (6, 18), playing an essential role in connecting the relaxosome complex with the secretion channel. The structural similarity between TrwB and well-known molecular motors, such as F 1 ATPase or ring helicases, suggests that TrwB uses the energy released from ATP hydrolysis to pump DNA through its central channel (19).Interactions between VirB11, VirB4, and VirD4 were mainly inferred from two-hybrid and coimmunoprecipitation assays (20)(21)(22). Here, we report direct evidence for interactions of TrwD with TrwK and TrwB proteins, the VirB11, VirB4, and VirD4 homologs in the conjugative plasmid R388.…”

mentioning

confidence: 99%

Functional Interactions of VirB11 Traffic ATPases with VirB4 and VirD4 Molecular Motors in Type IV Secretion Systems

Ripoll‐Rozada

Zunzunegui

Cruz

et al. 2013

Journal of Bacteriology

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Pilus biogenesis and substrate transport by type IV secretion systems require energy, which is provided by three molecular motors localized at the base of the secretion channel. One of these motors, VirB11, belongs to the superfamily of traffic ATPases, which includes members of the type II secretion system and the type IV pilus and archaeal flagellar assembly apparatus. Here, we report the functional interactions between TrwD, the VirB11 homolog of the conjugative plasmid R388, and TrwK and TrwB, the motors involved in pilus biogenesis and DNA transport, respectively. Although these interactions remained standing upon replacement of the traffic ATPase by a homolog from a phylogenetically related conjugative system, namely, TraG of plasmid pKM101, this homolog could not replace the TrwD function for DNA transfer. This result suggests that VirB11 works as a switch between pilus biogenesis and DNA transport and reinforces a mechanistic model in which VirB11 proteins act as traffic ATPases by regulating both events in type IV secretion systems. Type IV secretion systems (T4SSs) export proteins and virulence effectors to other bacteria and eukaryotic cells (1-3). They are also responsible for genetic exchange between bacteria during conjugation (4, 5). T4SSs are large macromolecular assemblies formed by 12 different protein subunits, named VirB1 to VirB11 and VirD4, following the Agrobacterium tumefaciens T4SS nomenclature. Three of these proteins (VirD4, VirB4, and VirB11) are hexameric ATPases (6-8) that provide the energy for substrate transport and T4SS biogenesis.VirB11 proteins are traffic ATPases that belong to a large AAA ϩ secretion protein superfamily, which also includes proteins of type II secretion systems and the type IV pilus biogenesis and archaeal flagellar assembly machineries (9). All of them are soluble, hexameric proteins located at the cytoplasmic side of the inner membrane. The monomer is characterized by the presence of an N-terminal domain (NTD) and a C-terminal domain (CTD), which are connected by a flexible linker that plays a key role in the catalytic cycle (10-12). Comparison of the crystal structures of VirB11 from Brucella suis (13) and its homolog in Helicobacter pylori, HP0525 (10,14), revealed that this linker is responsible for a large domain swap of the NTD over the CTD without affecting the hexameric assembly (13).VirB11 was reported to assist VirB4 during pilus biogenesis by dislocating pilin subunits from the inner membrane to the periplasmic space, thus promoting pilus polymerization (15). VirB4 proteins are the largest and most conserved components of T4SSs. TrwK, the VirB4 homolog of plasmid R388, consists of a hexameric double ring with a barrel-shaped structure (16). The atomic structure of the C-terminal domain of the VirB4 homolog in Thermoanaerobacter pseudethanolicus was recently obtained (17), revealing a striking structural similarity with TrwB, the VirD4 homolog of the R388 plasmid. TrwB was extensively characterized as a DNA-dependent ATPase (6, 18), playing an ess...

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Topology of the VirB4 C Terminus in the Agrobacterium tumefaciens VirB/D4 Type IV Secretion System

Cited by 23 publications

References 42 publications

ATPase Activity and Oligomeric State of TrwK, the VirB4 Homologue of the Plasmid R388 Type IV Secretion System

ATPase Activity and Oligomeric State of TrwK, the VirB4 Homologue of the Plasmid R388 Type IV Secretion System

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

Functional Interactions of VirB11 Traffic ATPases with VirB4 and VirD4 Molecular Motors in Type IV Secretion Systems

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