Formation of a β-barrel membrane protein is catalyzed by the interior surface of the assembly machine protein BamA

Lee, James; Tomasek, David; Santos, Thiago M. A.; May, Mary; Meuskens, Ina; Kahne, Daniel

doi:10.7554/elife.49787

Cited by 56 publications

(98 citation statements)

References 66 publications

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“…Growing evidence points to a mechanism of BAM function in catalyzing OMP folding and membrane insertion that involves the templating of C-terminal β-strands of the nascent substrate OMPs onto the β1 strand of BamA ( Figs. 6 – 8 ) ( 47 , 61 , 384 ). The C-terminal strands of OMPs in bacteria and mitochondria contain a conserved aromatic-rich motif termed the β-signal ( Fig.…”

Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

“…A conformational change in BAM then inserts the already-folded β-barrel (the swing/elongation models) ( 47 , 389 ). 4) Substrates fold against the interior wall of BamA while keeping their N and C termini in close proximity ready for β-barrel closure and release into the membrane (the lumen-catalyzed model) ( 61 ). The last two models are particularly intriguing as they suggest that the hydrophobic surface of the folding OMP is partially exposed to an aqueous or polar environment.…”

Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

“…The last two models are particularly intriguing as they suggest that the hydrophobic surface of the folding OMP is partially exposed to an aqueous or polar environment. How this step could be energetically favorable is not yet clear, but some authors have proposed that the cradle created by the BamA lumen and POTRA domains may aid folding by acting like an entropic cage, analogous to chaperonins such as GroEL/ES ( 61 , 389 ). This may drive folding by reducing the conformational entropy of the unfolded state but may also restrict the mobility of water, perhaps offsetting some of the cost of exposing hydrophobic residues.…”

Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

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Role of the lipid bilayer in outer membrane protein folding in Gram-negative bacteria

Horne

Brockwell

Radford

2020

Journal of Biological Chemistry

124

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β-barrel outer membrane proteins (OMPs) represent the major proteinaceous component of the outer membrane (OM) of Gram-negative bacteria. These proteins perform key roles in cell structure and morphology, nutrient acquisition, colonisation and invasion, and protection against external toxic threats such as antibiotics. To become functional, OMPs must fold and insert into a crowded and asymmetric OM that lacks much freely accessible lipid. This feat is accomplished in the absence of an external energy source and is thought to be driven by the high thermodynamic stability of folded OMPs in the OM. With such a stable fold, the challenge that bacteria face in assembling OMPs into the OM is how to overcome the initial energy barrier of membrane insertion. In this review, we highlight the roles of the lipid environment and the OM in modulating the OMP folding landscape and discuss the factors that guide folding in vitro and in vivo. We particularly focus on the composition, architecture and physical properties of the OM and how an understanding of the folding properties of OMPs in vitro can help explain the challenges they encounter during folding in vivo. Current models of OMP biogenesis in the cellular environment are still in flux, but the stakes for improving the accuracy of these models are high. Since OMP folding is an essential process in all Gram-negative bacteria, and considering the looming crisis of widespread microbial drug resistance, to bring down the powerful, OMP-supported barrier against antibiotics, we must first understand how bacterial cells build it.

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Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

Section: Bam: Nature's Answer To the Challenges Of Omp Foldingmentioning

confidence: 99%

See 1 more Smart Citation

Role of the lipid bilayer in outer membrane protein folding in Gram-negative bacteria

Horne

Brockwell

Radford

2020

Journal of Biological Chemistry

124

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“…Stabilisation of the seam by disulfide cross-linking 19 , or extension of one of the β-strands that form it 20 , is lethal in vivo, suggesting that opening of the lateral gate is essential for BAM function, at least for some substrate OMPs, in the context of a living bacterium. In support of the importance of these lateral gate dynamics, the N-terminal strand of the BamA (β1) barrel has been shown to hydrogen-bond to the substrate OMP, possibly via the β-signal in the C-terminal β-strand in both BAM 24,25 and its mitochondrial homologue, SAM 26 . The POTRA domains of BamA are mobile 23,27,28 and have been observed in various conformations in structures of BamABCDE [13][14][15] and BamACDE 12 , and this mobility is thought to be important for function, as reducing the flexibility between POTRA domains 2 and 3, by introducing a disulfide bond between them, also impairs BAM function in vivo 23 .…”

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confidence: 99%

Distortion of the bilayer and dynamics of the BAM complex in lipid nanodiscs

et al. 2020

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The β-barrel assembly machinery (BAM) catalyses the folding and insertion of β-barrel outer membrane proteins (OMPs) into the outer membranes of Gram-negative bacteria by mechanisms that remain unclear. Here, we present an ensemble of cryoEM structures of the E. coli BamABCDE (BAM) complex in lipid nanodiscs, determined using multi-body refinement techniques. These structures, supported by single-molecule FRET measurements, describe a range of motions in the BAM complex, mostly localised within the periplasmic region of the major subunit BamA. The β-barrel domain of BamA is in a ‘lateral open’ conformation in all of the determined structures, suggesting that this is the most energetically favourable species in this bilayer. Strikingly, the BAM-containing lipid nanodisc is deformed, especially around BAM’s lateral gate. This distortion is also captured in molecular dynamics simulations, and provides direct structural evidence for the lipid ‘disruptase’ activity of BAM, suggested to be an important part of its functional mechanism.

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“…The SAM complex consists of three hairpin (or larger portion) of the substrate enters the BamA lumen, associates with unpaired bstrands on BamA, and uses the BamA b-barrel to sequentially fold into the outer membrane, budding off from BamA when the first and last strands of the substrate come together. With the wide variety and large quantities of b-barrel proteins that exist in bacteria, it is possible that either or both models, or variations of these 20,21 , are used depending on the complexity of the substrate.…”

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confidence: 99%

Structural Insight into Mitochondrial β-Barrel Outer Membrane Protein Biogenesis

Diederichs

Rollauer

et al. 2020

Preprint

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In mitochondria, b-barrel outer membrane proteins mediate protein import, metabolite transport, lipid transport, and biogenesis. The Sorting and Assembly Machinery (SAM) complex consists of three proteins that assemble as a 1:1:1 complex to fold b-barrel proteins and insert them into the mitochondrial outer membrane. We report cryoEM structures of the SAM complex from Myceliophthora thermophila, which show that Sam50 forms a 16-stranded transmembrane b-barrel with a single polypeptide-transport-associated (POTRA) domain extending into the intermembrane space. Sam35 and Sam37 are located on the cytosolic side of the outer membrane, with Sam35 capping Sam50, and Sam37 interacting extensively with Sam35. Sam35 and Sam37 each adopt a GST-like fold, with no functional, structural, or sequence similarity to their bacterial counterparts. Structural analysis shows how the Sam50 bbarrel opens a lateral gate to accommodate its substrates. The SAM complex structure suggests how it interacts with other mitochondrial outer membrane proteins to create supercomplexes. components: a b-barrel core, Sam50, which spans the outer membrane, and two accessory subunits, Sam35 and Sam37, that associate with Sam50 on the cytosolic side of the membrane 2-5 . Sam50 and Sam35 are essential proteins, with Sam50 folding and inserting b-barrel substrates into the outer membrane and Sam35 interacting with the substrate b-signal located in the last b-strand 3 . Sam37, while not essential, functions in substrate release 3,6 and may also promote formation of a TOM-SAM supercomplex 7 .Bacterial b-barrel membrane proteins take a different pathway to reach the outer membrane, but they are inserted by evolutionarily related machinery 8 . In bacteria, proteins are synthesized in the cytoplasm, secreted across the inner membrane by the Sec translocon, bound to chaperones in the periplasm, and transferred to the Bacterial Assembly Machinery (BAM) complex for folding and insertion into the outer membrane 9,10 . In Escherichia coli, four lipoproteins, BamB, BamC, BamD, and BamE, associate with the periplasmic domain of BamA (itself a 16-stranded transmembrane b-barrel) to fold and insert b-barrels ranging in size and complexity from 8 b-strands with a simple barrel fold, to 26 b-strands with multiple domains 11 . Structures of BamA and BAM complexes illustrate two features of BamA that facilitate folding and insertion: [1] BamA has a narrowed hydrophobic surface where the first and last b-strands meet, which locally compresses and destabilizes the lipid bilayer to allow membrane protein insertion (BamA assisted model). [2] The first and last b-strands form a 'lateral gate' that has been shown to open and close by molecular dynamics (MD) simulations, disulfide crosslinking, and structures solved by X-ray crystallography and cryo-electron microscopy (cryoEM) 12-19 . The required opening and closing of the lateral gate gave rise to the budding model, where a b-

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Formation of a β-barrel membrane protein is catalyzed by the interior surface of the assembly machine protein BamA

Cited by 56 publications

References 66 publications

Role of the lipid bilayer in outer membrane protein folding in Gram-negative bacteria

Role of the lipid bilayer in outer membrane protein folding in Gram-negative bacteria

Distortion of the bilayer and dynamics of the BAM complex in lipid nanodiscs

Structural Insight into Mitochondrial β-Barrel Outer Membrane Protein Biogenesis

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