BamE Modulates the Escherichia coli Beta-Barrel Assembly Machine Component BamA

Rigel, Nathan W.; Schwalm, Jaclyn; Ricci, Dante P.; Silhavy, Thomas J.

doi:10.1128/jb.06426-11

Cited by 80 publications

(91 citation statements)

References 30 publications

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“…In previous studies, we uncovered evidence that both BamA and BamD can exist in different states. In contrast to the protease-resistant BamA form found in wild-type cells, BamA exists primarily in an alternate, protease-sensitive conformation in the absence of BamE (8). Additionally, we isolated a gainof-function mutation in bamD (bamDR197L) that was proposed to stabilize the activated state of BamD (12).…”

Section: Resultsmentioning

confidence: 99%

“…Single bam mutations that cause subtle defects often exhibit synergistic effects when combined (5,8). As these phenotypes have the potential to both unmask critical roles for the various components in OMP assembly and reveal the step(s) at which they act, we sought to determine whether such interactions are observed when accessory Bam components are deleted in a bamAE373A mutant background.…”

Section: Deletion Of Accessory Bam Genes Causes Lethality In a Bamae373amentioning

confidence: 99%

“…We have shown that BamE functions uniquely among the nonessential Bam lipoproteins; specifically, BamE modulates the conformation of the BamA β-barrel domain (8). Because BamE is not thought to directly interact with BamA, this activity is presumably mediated through BamD.…”

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confidence: 99%

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Conformation-specific labeling of BamA and suppressor analysis suggest a cyclic mechanism for β-barrel assembly in Escherichia coli

Rigel

Ricci

Silhavy

2013

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

120

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In Gram-negative bacteria, integral outer membrane β-barrel proteins (OMPs) are assembled by the beta-barrel assembly machine (Bam) complex. The essential components of this complex are the OMP BamA [which contains a carboxyl-terminal β-barrel and an amino-terminal periplasmic module composed of five polypeptide transport associated (POTRA) domains] and the lipoprotein BamD. In Escherichia coli, the Bam complex also contains three nonessential lipoproteins (BamBCE), all of which require the barrel-proximal POTRA domain (P5) for stable interactions with BamA. We have previously reported that the BamA β-barrel assumes two different conformations. A method for conformation-specific labeling of BamA described here reveals that these conformers reflect the degree of surface exposure of the conserved sixth extracellular loop (L6). L6 is surface accessible in one conformation but not in the other, likely because it occupies the lumen of the BamA β-barrel in the latter case. A gain-of-function mutation that promotes Bam activity (bamDR197L) and a loss-of-function mutation that decreases the activity of Bam (ΔbamE) both favor surface exposure of BamA L6, suggesting that BamD and BamE normally act to control L6 exposure through opposing functions. These results, along with the synthetic lethality of the bamDR197L ΔbamE double mutant, imply a cyclic mechanism in which the Bam lipoproteins regulate the conformation of BamA during the OMP assembly reaction. Our results further suggest that BamDE controls L6 exposure via conformational signals transmitted through P5 to L6.genetics | OM biogenesis | Omp85 | membrane protein folding |

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Section: Resultsmentioning

confidence: 99%

Section: Deletion Of Accessory Bam Genes Causes Lethality In a Bamae373amentioning

confidence: 99%

See 1 more Smart Citation

Conformation-specific labeling of BamA and suppressor analysis suggest a cyclic mechanism for β-barrel assembly in Escherichia coli

Rigel

Ricci

Silhavy

2013

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

120

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“…Collectively, these results suggest that proper assembly of BamA is compromised in the absence of SurA and that BepA degrades BamA under such a condition. Alternatively, as BamA has been suggested to assume different conformations during its functional cycle (45,46), the absence of SurA might cause accumulation of the BepA-susceptible form of BamA.…”

Section: Bepa Degrades Lptd When Lptd Fails To Form the Om Lps Translmentioning

confidence: 99%

Protease homolog BepA (YfgC) promotes assembly and degradation of β-barrel membrane proteins in Escherichia coli

Narita

Masui

Suzuki

et al. 2013

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

135

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Gram-negative bacteria are equipped with quality-control systems for the outer membrane (OM) that sense and cope with defective biogenesis of its components. Accumulation of misfolded outer membrane proteins (OMPs) in Escherichia coli leads to activation of σ E , an essential alternative σ factor that up-regulates transcription of multiple genes required to preserve OM structure and function. Disruption of bepA (formerly yfgC), a σ E -regulated gene encoding a putative periplasmic metalloprotease, sensitizes cells to multiple drugs, suggesting that it may be involved in maintaining OM integrity. However, the specific function of BepA remains unclear. Here, we show that BepA enhances biogenesis of LptD, an essential OMP involved in OM transport and assembly of lipopolysaccharide, by promoting rearrangement of intramolecular disulfide bonds of LptD. In addition, BepA possesses protease activity and is responsible for the degradation of incorrectly folded LptD. In the absence of periplasmic chaperone SurA, BepA also promotes degradation of BamA, the central OMP subunit of the β-barrel assembly machinery (BAM) complex. Interestingly, defective oxidative folding of LptD caused by bepA disruption was partially suppressed by expression of protease-active site mutants of BepA, suggesting that BepA functions independently of its protease activity. We also show that BepA has genetic and physical interaction with components of the BAM complex. These findings raised the possibility that BepA maintains the integrity of OM both by promoting assembly of OMPs and by proteolytically eliminating OMPs when their correct assembly was compromised.extracytoplasmic function sigma factor | protein quality control | disulfide bond formation | peptidase M48 | tetratricopeptide repeat (TPR) motif

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“…BamD is the only lipoprotein known to be essential for a functional BAM complex (Wu et al, 2005, Onufryk et al, 2005, Malinverni et al, 2006. BamE and BamC are thought to stabilise the interaction between BamA and BamD, and to modulate the functional activity of BamA and BamD, respectively (Kim et al, 2011a, Rigel et al, 2012. BamB is known to form a propeller like structure within the periplasm and is linked together with BamA via the third POTRA domain (Jansen et al, 2015a, Dong et al, 2012b.…”

Section: Growth Characteristics Of Mg1655 Periplasmic Chaperone Mutanmentioning

confidence: 99%

Autotransporter proteins of Uropathogenic Escherichia coli: regulation, characterisation and the role of periplasmic proteins in their expression

Nichols¹

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Uropathogenic Escherichia coli (UPEC) are the primary cause for all urinary tract infections.Autotransporter (AT) proteins are virulence factors secreted by the type V secretion pathway. This project examined the prevalence, regulation and expression of the vacuolating AT toxin (Vat), and the role of periplasmic chaperone proteins in AT translocation.The vacuolating autotransporter (AT) toxin (Vat) contributes to Uropathogenic Escherichia coli (UPEC) fitness during systemic infection. Vat is a serine protease AT of Enterobacteriaceae (SPATE) with cytotoxic activity. Here we characterised Vat and investigated its regulation in UPEC.We assessed the prevalence of vat in a collection of 45 UPEC urosepsis strains and showed it that was present in 31 (68%) of the isolates. The isolates containing the vat gene corresponded to three major E. coli sequence types (ST12, 73 and 95) and these strains secreted the Vat protein.Further analysis of the vat genetic locus identified a conserved gene located directly downstream of vat that encodes a putative MarR-like transcriptional regulator, which we termed vatX. The vatvatX genes were present in the UPEC reference strain CFT073 and RT-PCR revealed both genes are co-transcribed. Over-expression of vatX in CFT073 led to a 3-fold increase in vat gene transcription. The vat promoter region contained three putative nucleation sites for the global transcriptional regulator H-NS; thus the hns gene was mutated in CFT073 (to generate CFT073hns).Western blot analysis using a Vat-specific antibody revealed a significant increase in Vat expression in CFT073hns compared to wild-type CFT073. Direct H-NS binding to the vat promoter region was demonstrated using purified H-NS in combination with electrophoresis mobility shift assays. Finally, Vat-specific antibodies were detected in plasma samples from urosepsis patients iv Declaration by authorThis thesis is composed of my original work, and contains no material previously published or written by another person except where due reference has been made in the text. I have clearly stated the contribution by others to jointly-authored works that I have included in my thesis.

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BamE Modulates the Escherichia coli Beta-Barrel Assembly Machine Component BamA

Cited by 80 publications

References 30 publications

Conformation-specific labeling of BamA and suppressor analysis suggest a cyclic mechanism for β-barrel assembly in Escherichia coli

Conformation-specific labeling of BamA and suppressor analysis suggest a cyclic mechanism for β-barrel assembly in Escherichia coli

Protease homolog BepA (YfgC) promotes assembly and degradation of β-barrel membrane proteins in Escherichia coli

Autotransporter proteins of Uropathogenic Escherichia coli: regulation, characterisation and the role of periplasmic proteins in their expression

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