Structural insight into the biogenesis of β-barrel membrane proteins

Noinaj, Nicholas; Kuszak, Adam; Gumbart, James C.; Lukacik, P.; Chang, Hoshing; Easley, N.; Lithgow, Trevor; Buchanan, Susan K.

doi:10.1038/nature12521

Cited by 393 publications

(727 citation statements)

References 62 publications

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“…Although structures have been reported for the orthologs of the Omp85 family, including FhaC, TamA, and BamA, in Gram-negative bacteria (35,36,(43)(44)(45)(46), no structure has been reported for either Sam50 in mitochondria or Toc75 or OEP80 in chloroplasts. Here we report the structure of Toc75, from A. thaliana, consisting of the N-terminal linker and three tandem POTRA domains (Figs.…”

Section: Discussionmentioning

confidence: 99%

“…The structure reveals that the POTRA domains form an L-shaped structure very similar to BamA, with a unique extended helix within POTRA2 that is not observed in the POTRA domains of BamA and other Omp85 family members (25,28,(35)(36)(37)(38). Based on these unique structural properties, we investigated the contribution of individual POTRA repeats in binding chloroplast precursors.…”

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confidence: 99%

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The POTRA domains of Toc75 exhibit chaperone-like function to facilitate import into chloroplasts

O'Neil

Richardson

Paila

et al. 2017

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

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Protein trafficking across membranes is an essential function in cells; however, the exact mechanism for how this occurs is not well understood. In the endosymbionts, mitochondria and chloroplasts, the vast majority of proteins are synthesized in the cytoplasm as preproteins and then imported into the organelles via specialized machineries. In chloroplasts, protein import is accomplished by the TOC (translocon on the outer chloroplast membrane) and TIC (translocon on the inner chloroplast membrane) machineries in the outer and inner envelope membranes, respectively. TOC mediates initial recognition of preproteins at the outer membrane and includes a core membrane channel, Toc75, and two receptor proteins, Toc33/34 and Toc159, each containing GTPase domains that control preprotein binding and translocation. Toc75 is predicted to have a β-barrel fold consisting of an N-terminal intermembrane space (IMS) domain and a C-terminal 16-stranded β-barrel domain. Here we report the crystal structure of the N-terminal IMS domain of Toc75 from Arabidopsis thaliana, revealing three tandem polypeptide transportassociated (POTRA) domains, with POTRA2 containing an additional elongated helix not observed previously in other POTRA domains. Functional studies show an interaction with the preprotein, preSSU, which is mediated through POTRA2-3. POTRA2-3 also was found to have chaperone-like activity in an insulin aggregation assay, which we propose facilitates preprotein import. Our data suggest a model in which the POTRA domains serve as a binding site for the preprotein as it emerges from the Toc75 channel and provide a chaperonelike activity to prevent misfolding or aggregation as the preprotein traverses the intermembrane space.protein import | chloroplast | outer membrane | Toc75 | POTRA domain

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

The POTRA domains of Toc75 exhibit chaperone-like function to facilitate import into chloroplasts

O'Neil

Richardson

Paila

et al. 2017

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

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“…Thus, strict pore gating is critical for bacterial homeostasis. Other β-barrel proteins that shuttle constituents across or into the outer membrane use the energy of ATP or the membrane potential across the inner membrane to regulate a gated channel, for instance the TonB-dependent translocators (29), or as in the case of the recently solved BamA, seem to facilitate insertion of proteins into the outer membrane by opening/closing of the BamA β-barrel (30). In CUP ushers, OM integrity is maintained via the existence of a β-sandwich PLUG domain, which resides in the transmembrane channel in the absence of pilus assembly (Fig.…”

Section: Discussionmentioning

confidence: 99%

Molecular basis of usher pore gating in Escherichia coli pilus biogenesis

Volkan

Kalas

Pinkner³

et al. 2013

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

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Extracellular fibers called chaperone-usher pathway pili are critical virulence factors in a wide range of Gram-negative pathogenic bacteria that facilitate binding and invasion into host tissues and mediate biofilm formation. Chaperone-usher pathway ushers, which catalyze pilus assembly, contain five functional domains: a 24-stranded transmembrane β-barrel translocation domain (TD), a β-sandwich plug domain (PLUG), an N-terminal periplasmic domain, and two Cterminal periplasmic domains (CTD1 and 2). Pore gating occurs by a mechanism whereby the PLUG resides stably within the TD pore when the usher is inactive and then upon activation is translocated into the periplasmic space, where it functions in pilus assembly. Using antibiotic sensitivity and electrophysiology experiments, a single salt bridge was shown to function in maintaining the PLUG in the TD channel of the P pilus usher PapC, and a loop between the 12th and 13th beta strands of the TD (β12-13 loop) was found to facilitate pore opening. Mutation of the β12-13 loop resulted in a closed PapC pore, which was unable to efficiently mediate pilus assembly. Deletion of the PapH terminator/anchor resulted in increased OM permeability, suggesting a role for the proper anchoring of pili in retaining OM integrity. Further, we introduced cysteine residues in the PLUG and N-terminal periplasmic domains that resulted in a FimD usher with a greater propensity to exist in an open conformation, resulting in increased OM permeability but no loss in type 1 pilus assembly. These studies provide insights into the molecular basis of usher pore gating and its roles in pilus biogenesis and OM permeability.outer membrane usher | macromolecular assembly | bacterial pathogenesis

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“…Lipids near the b1-b16 strands are highlighted with carbons C1, C2 and C3 coloured with blue spheres; lipids near the opposite side of the BamA b-barrel are highlighted with orange spheres. The lipid atom selection was carried out as described by Noinaj et al [40].…”

Section: Introduction: the Challenges Of Unfolded Outer Membrane Protmentioning

confidence: 99%

A combined kinetic push and thermodynamic pull as driving forces for outer membrane protein sorting and folding in bacteria

Fleming

2015

Phil. Trans. R. Soc. B

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One contribution of 12 to a theme issue 'The bacterial cell envelope'. In vitro folding studies of outer membrane beta-barrels have been invaluable in revealing the lipid effects on folding rates and efficiencies as well as folding free energies. Here, the biophysical results are summarized, and these kinetic and thermodynamic findings are considered in terms of the requirements for folding in the context of the cellular environment. Because the periplasm lacks an external energy source the only driving forces for sorting and folding available within this compartment are binding or folding free energies and their associated rates. These values define functions for periplasmic chaperones and suggest a biophysical mechanism for the BAM complex.

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Structural insight into the biogenesis of β-barrel membrane proteins

Cited by 393 publications

References 62 publications

The POTRA domains of Toc75 exhibit chaperone-like function to facilitate import into chloroplasts

The POTRA domains of Toc75 exhibit chaperone-like function to facilitate import into chloroplasts

Molecular basis of usher pore gating in Escherichia coli pilus biogenesis

A combined kinetic push and thermodynamic pull as driving forces for outer membrane protein sorting and folding in bacteria

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