Analysis of YfgL and YaeT Interactions through Bioinformatics, Mutagenesis, and Biochemistry

Vuong, Phu; Bennion, Drew; Mantei, J.; Frost, Danielle; Misra, Rajeev

doi:10.1128/jb.01477-07

Cited by 93 publications

(159 citation statements)

References 44 publications

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“…Considering its relatively low abundance in the eluates and the fact that BamB did not appear to be required for Hbp secretion, its copurification with the Hbp110C/348C intermediate may reflect secondary cross-linking to BamA rather than a direct interaction with the intermediate itself. In any case, a close proximity of BamB is likely, since a direct interaction between BamA and BamB has been demonstrated recently (Vuong et al, 2008). The copurification of OmpA is more enigmatic since it is not a known constituent of the Bam complex.…”

Section: Discussionmentioning

confidence: 99%

The Bam (Omp85) complex is involved in secretion of the autotransporter haemoglobin protease

et al. 2009

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Autotransporters are large virulence factors secreted by Gram-negative bacteria. They are synthesized with a C-terminal domain that forms a b-barrel pore in the outer membrane implicated in translocation of the upstream 'passenger' domain across the outer membrane. However, recent structural data suggest that the diameter of the b-barrel pore is not sufficient to allow the passage of partly folded structures observed for several autotransporters. Here, we have used a stalled translocation intermediate of the autotransporter Hbp to identify components involved in insertion and translocation of the protein across the outer membrane. At this intermediate stage the b-domain was not inserted and folded as an integral b-barrel in the outer membrane whereas part of the passenger was surface exposed. The intermediate was copurified with the periplasmic chaperone SurA and subunits of the Bam (Omp85) complex that catalyse the insertion and assembly of outer-membrane proteins. The data suggest a critical role for this general machinery in the translocation of autotransporters across the outer membrane.

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Section: Discussionmentioning

confidence: 99%

The Bam (Omp85) complex is involved in secretion of the autotransporter haemoglobin protease

et al. 2009

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“…In E. coli, BamC, BamD and BamE interact with BamA as a group, while the BamA-BamB interaction is independent of the other lipoproteins (Malinverni et al, 2006;Sklar et al, 2007;Vuong et al, 2008). In mutants lacking either BamC or BamE, BamD levels are normal but less BamD is associated with BamA.…”

Section: Discussionmentioning

confidence: 99%

“…A previous study of the E. coli BAM complex identified alleles of bamB (yfgL) that caused little or no phenotypic defect, yet weakened the interaction between BamB and BamA in co-immunoprecipitation experiments, suggesting that the affected amino acids are important for BAM complex stability (Vuong et al, 2008). To generate intermediate defects in chemical sensitivity (Table 2), either the function of Caulobacter BamEDC or the stability of the resulting BAM complex could be compromised.…”

Section: Under Conditions Of Phosphate Limitation Wild-typementioning

confidence: 99%

“…Within the BAM complex, BamB interacts directly with BamA, while BamC, D and E depend on each other for interaction with BamA (Malinverni et al, 2006;Sklar et al, 2007;Vuong et al, 2008). Because Caulobacter lacks a BamC homologue (Gatsos et al, 2008), we wondered if BamE (and BamD) could still associate with BamA.…”

Section: Caulobacter Bame Associates With Other Proteins Implicated Imentioning

confidence: 99%

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The BAM complex subunit BamE (SmpA) is required for membrane integrity, stalk growth and normal levels of outer membrane β-barrel proteins in Caulobacter crescentus

2010

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The outer membrane of Gram-negative bacteria is an essential compartment containing a specific complement of lipids and proteins that constitute a protective, selective permeability barrier. Outer membrane β-barrel proteins are assembled into the membrane by the essential hetero-oligomeric BAM complex, which contains the lipoprotein BamE. We have identified a homologue of BamE, encoded by CC1365, which is located in the outer membrane of the stalked alpha-proteobacterium Caulobacter crescentus. BamE associates with proteins whose homologues in other bacteria are known to participate in outer membrane protein assembly: BamA (CC1915), BamB (CC1653) and BamD (CC1984). Caulobacter cells lacking BamE grow slowly in rich medium and are hypersensitive to anionic detergents, some antibiotics and heat exposure, which suggest that the membrane integrity of the mutant is compromised. Membranes of the ΔbamE mutant have normal amounts of the outer membrane protein RsaF, a TolC homologue, but are deficient in CpaC*, an aggregated form of the outer membrane secretin for type IV pili. ΔbamE membranes also contain greatly reduced amounts of three TonB-dependent receptors that are abundant in wild-type cells. Cells lacking BamE have short stalks and are delayed in stalk outgrowth during the cell cycle. Based on these findings, we propose that Caulobacter BamE participates in the assembly of outer membrane β-barrel proteins, including one or more substrates required for the initiation of stalk biogenesis.

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“…Many in vitro and in vivo studies have investigated various components involved in this pathway (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). However, in vitro experiments typically characterize individual components in isolation, and in vivo studies are often incapable of deciphering how specific components are responsible for observed phenomena.…”

mentioning

confidence: 99%

Dynamic periplasmic chaperone reservoir facilitates biogenesis of outer membrane proteins

Costello

Plummer

Fleming

2016

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

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Outer membrane protein (OMP) biogenesis is critical to bacterial physiology because the cellular envelope is vital to bacterial pathogenesis and antibiotic resistance. The process of OMP biogenesis has been studied in vivo, and each of its components has been studied in isolation in vitro. This work integrates parameters and observations from both in vivo and in vitro experiments into a holistic computational model termed "Outer Membrane Protein Biogenesis Model" (OMPBioM). We use OMPBioM to assess OMP biogenesis mathematically in a global manner. Using deterministic and stochastic methods, we are able to simulate OMP biogenesis under varying genetic conditions, each of which successfully replicates experimental observations. We observe that OMPs have a prolonged lifetime in the periplasm where an unfolded OMP makes, on average, hundreds of short-lived interactions with chaperones before folding into its native state. We find that some periplasmic chaperones function primarily as quality-control factors; this function complements the folding catalysis function of other chaperones. Additionally, the effective rate for the β-barrel assembly machinery complex necessary for physiological folding was found to be higher than has currently been observed in vitro. Overall, we find a finely tuned balance between thermodynamic and kinetic parameters maximizes OMP folding flux and minimizes aggregation and unnecessary degradation. In sum, OMPBioM provides a global view of OMP biogenesis that yields unique insights into this essential pathway.T he cellular envelope of Gram-negative bacteria is comprised of two membranes separated by an aqueous compartment termed the "periplasm." The outer membrane of the cellular envelope contains integral β-barrel membrane proteins referred to as "outer membrane proteins" (OMPs) (1, 2). The outer membrane and OMPs provide the first barrier between bacteria and the environment and are essential to many important cellular processes including metabolic transport, bacterial virulence, and antibiotic resistance (3-5). Understanding the pathway by which OMPs traverse the periplasm and attain their native functional state is essential to an ability to manipulate this element of the bacterial cell.The OMP biogenesis process is distinct from the folding of cytosolic proteins because it involves a unique collection of obstacles. First, OMPs do not adopt their folded conformations while in an aqueous environment (6). Rather, unfolded OMPs (uOMPs) must be transported across the periplasm to reach their native membrane. Because of their marginal solubility in water, this process must be tightly controlled to avoid aggregation. Second, structures of folded OMPs (fOMPs) show that these proteins contain water-solvated residues in loops on the outer surfaces of bacteria. The desolvation and transport of these polar and ionizable side chains across the outer membrane represent significant kinetic barriers to OMP folding (7). Third, Gram-negative bacteria maintain a high density of fOMP in the expanding oute...

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Analysis of YfgL and YaeT Interactions through Bioinformatics, Mutagenesis, and Biochemistry

Cited by 93 publications

References 44 publications

The Bam (Omp85) complex is involved in secretion of the autotransporter haemoglobin protease

The Bam (Omp85) complex is involved in secretion of the autotransporter haemoglobin protease

The BAM complex subunit BamE (SmpA) is required for membrane integrity, stalk growth and normal levels of outer membrane β-barrel proteins in Caulobacter crescentus

Dynamic periplasmic chaperone reservoir facilitates biogenesis of outer membrane proteins

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