Plasticity within the barrel domain of BamA mediates a hybrid-barrel mechanism by BAM

Wu, Runrun; Bakelar, Jeremy; Lundquist, Karl; Zhang, Zijian; Kuo, Katie M.; Ryoo, David; Pang, Yui Tik; Sun, Chen; White, Tommi; Klose, Thomas; Jiang, Wen; Gumbart, James C.; Noinaj, Nicholas

doi:10.1038/s41467-021-27449-4

Cited by 52 publications

(51 citation statements)

References 88 publications

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“…In vitro and in silico evidence point to a role for BAM in membrane thinning and disordering of the OM bilayer to aid OMP folding 46 – 50 . Models in which the substrate makes direct interactions with the BamA barrel have also been proposed 5 , 6 , 45 , 51 – 53 , and are supported by a variety of experimental evidence 54 – 56 , including in vivo crosslinking 53 , 57 , 58 and structural data 59 – 62 . For example, a cryoEM structure of a late-stage folding intermediate of BamA being folded by BAM 59 , as well as recent structures of EspP stalled while folding on BAM 61 , 62 , indicate that the terminal strand of an incoming OMP makes a β-augmentation interaction with the β1 strand of the BamA barrel as part of the assembly mechanism.…”

Section: Introductionmentioning

confidence: 83%

“…Models in which the substrate makes direct interactions with the BamA barrel have also been proposed 5 , 6 , 45 , 51 – 53 , and are supported by a variety of experimental evidence 54 – 56 , including in vivo crosslinking 53 , 57 , 58 and structural data 59 – 62 . For example, a cryoEM structure of a late-stage folding intermediate of BamA being folded by BAM 59 , as well as recent structures of EspP stalled while folding on BAM 61 , 62 , indicate that the terminal strand of an incoming OMP makes a β-augmentation interaction with the β1 strand of the BamA barrel as part of the assembly mechanism. Additional structures of BamA/BAM in which OMP-derived β-strands 60 , 63 or darobactin (a peptide antibiotic that targets BamA) 64 , 65 , are bound to β1 of the BamA barrel suggest that the lateral-closed state may be responsible for the receipt of unfolded OMPs for folding into the OM.…”

Section: Introductionmentioning

confidence: 83%

“…Here, we have shown that protection from HDX occurs in the BamA barrel in the presence of SurA, suggesting that binding of the chaperone to the periplasmic region of BAM may favour the BamA lateral-closed state. Recent structural and simulation evidence suggest that the lateral-open conformation of BAM is the major populated state in the absence of SurA 62 . Thus, we propose a model in which SurA acts to ready the BAM machinery for OMP folding by modulating the BAM conformational ensemble to favour the acceptor state.…”

Section: Discussionmentioning

confidence: 99%

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Dynamic interplay between the periplasmic chaperone SurA and the BAM complex in outer membrane protein folding

et al. 2022

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Correct folding of outer membrane proteins (OMPs) into the outer membrane of Gram-negative bacteria depends on delivery of unfolded OMPs to the β-barrel assembly machinery (BAM). How unfolded substrates are presented to BAM remains elusive, but the major OMP chaperone SurA is proposed to play a key role. Here, we have used hydrogen deuterium exchange mass spectrometry (HDX-MS), crosslinking, in vitro folding and binding assays and computational modelling to show that the core domain of SurA and one of its two PPIase domains are key to the SurA-BAM interaction and are required for maximal catalysis of OMP folding. We reveal that binding causes changes in BAM and SurA conformation and/or dynamics distal to the sites of binding, including at the BamA β1-β16 seam. We propose a model for OMP biogenesis in which SurA plays a crucial role in OMP delivery and primes BAM to accept substrates for folding.

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Section: Introductionmentioning

confidence: 83%

Section: Introductionmentioning

confidence: 83%

Section: Discussionmentioning

confidence: 99%

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Dynamic interplay between the periplasmic chaperone SurA and the BAM complex in outer membrane protein folding

et al. 2022

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“…The b5-b6 hairpin most likely binds by b augmentation of the barrel strand B1, by analogy with BamA and Sam50 in which the unzipped B1 strand templates folding of client proteins by b augmentation (Doyle & Bernstein, 2019;Höhr et al, 2018;Tomasek et al, 2020;Wu et al, 2021). This mode of binding is supported by our CIU results showing an increased cross-section of the protein upon binding of b5-b6 to the barrel and by the in vivo formation of an S-S bond between the tip of that hairpin and the extracellular loop L1.…”

Section: Discussionmentioning

confidence: 99%

Large-scale conformational changes of FhaC provide insights into the two-partner secretion mechanism

Sicoli

Konijnenberg

Guérin

et al. 2021

Preprint

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The Two-Partner secretion pathway mediates protein transport across the outer membrane of Gram-negative bacteria. TpsB transporters belong to the Omp85 superfamily, whose members catalyze protein insertion into, or translocation across membranes without external energy sources. They are composed of a transmembrane β barrel preceded by two periplasmic POTRA domains that bind the incoming protein substrate. Here we used an integrative approach combining in vivo assays, mass spectrometry, nuclear magnetic resonance and electron paramagnetic resonance techniques suitable to detect minor states in heterogeneous populations, to explore transient conformers of the TspB transporter FhaC. This revealed substantial, spontaneous conformational changes with a portion of the POTRA2 domain coming close to the lipid bilayer and surface loops. Specifically, the amphipathic β hairpin immediately preceding the first barrel strand can insert into the β barrel. We propose that these motions enlarge the channel and may hoist the substrate into it for secretion. An anchor region at the interface of the β barrel and the POTRA2 domain stabilizes the transporter in the course of secretion. Our data propose a solution to the conundrum how these proteins mediate protein secretion without the need for cofactors, by utilizing intrinsic protein dynamics.

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“…BamD is docked to the POTRA domains, contributing to a large, funnel-like periplasmic feature of the BAM complex in the periplasm [16][17][18]23 . Recent focus on the interaction between the lateral gate of BamA and the β-signal in substrate has been made possible through obtaining snap-shot structures of assembly intermediates bound to the lateral gate 9,12,27 as well as the isolation of inhibitory compounds such as darobactin capable of entering the lateral gate of BamA to exert a lethal effect on E. coli 10,28,29 . Despite these recent advances, the molecular mechanism of OMP assembly is enigmatic.…”

Section: Mainmentioning

confidence: 99%

Dual Recognition of Multiple Signals in Bacterial Outer Membrane Proteins Enhance Assembly and Maintain Membrane Integrity

Ding

Imai

Bamert

et al. 2021

Preprint

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SUMMARYGram-negative bacteria, mitochondria and chloroplasts contain outer membrane proteins (OMPs) characterized by a transmembrane domain with a β barrel structure. Most OMPs have a diagnostic “β-signal” imprinted in the amino acid sequence of the final β-strand (−1 strand) of the β barrel. Molecular chaperones of the Omp85 superfamily, such as BamA in the bacterial BAM complex, recognizes the β-signal and then catalyze the folding of the OMP into its membrane-embedded β barrel structure. Here, we reconstituted OMP assembly in vitro to study this process with five model OMPs, revealing that a critical assembly signal exists in the fifth β-strand from the C-terminus (−5 strand). This signal was shown to drive a critical insertion step of the OMP assembly process but is not required for the initial recognition step between the OMP and BamA. Furthermore, we identified the receptor for this “-5 signal” as BamD, a second essential subunit of the BAM complex. Distinct binding sites for the β-signal and the -5 signal, were identified on BamD. Comparative sequence analysis showed that the -5 signal is a conserved feature in bacterial OMPs, and that mitochondrial OMPs also contains -5 signal motif positioned in the fifth from last strand of their β barrel structure. We propose the “-5 rule” as a conserved feature of the process of OMP assembly in bacteria and the organelles of eukaryotes.HIGHLIGHTSGram-negative bacterial OMPs contain a conserved motif at the fifth β-strand from the C-terminus (−5 strand).The information encrypted in the conserved -5 signal is responsible for a rate-limiting step in protein insertion into the outer membraneDistinct binding sites on the outer membrane protein BamD recognize the -5 signal and a canonical β-signal.We propose -5 rule, to explain the function of the -5 signal in OMP assembly.

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Plasticity within the barrel domain of BamA mediates a hybrid-barrel mechanism by BAM

Cited by 52 publications

References 88 publications

Dynamic interplay between the periplasmic chaperone SurA and the BAM complex in outer membrane protein folding

Dynamic interplay between the periplasmic chaperone SurA and the BAM complex in outer membrane protein folding

Large-scale conformational changes of FhaC provide insights into the two-partner secretion mechanism

Dual Recognition of Multiple Signals in Bacterial Outer Membrane Proteins Enhance Assembly and Maintain Membrane Integrity

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