An alternative topological model for <i>Escherichia coli</i> OmpA

Stathopoulos, Christos

doi:10.1002/pro.5560050122

Cited by 41 publications

(39 citation statements)

References 39 publications

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“…Jeanteur et al [85] predicted two transmembrane β—strands within the C -terminal domain, and additional C -terminal transmembrane segments were predicted by algorithms of Schirmer and Cowan [86] and Ferenci [87]. Stathapoulos [88] constructed a 16 β-barrel structure with eight transmembrane segments in the C -terminal domain that is consistent with the varied biochemical, immunological, and genetic topological data concerning OmpA (Figure 8). …”

Section: The Role Of Cphb In Folding Of Outer Membrane Protein a (mentioning

confidence: 68%

The Role of Short-Chain Conjugated Poly-(R)-3-Hydroxybutyrate (cPHB) in Protein Folding

Reusch

2013

IJMS

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Poly-(R)-3-hydroxybutyrate (PHB), a linear polymer of R-3-hydroxybutyrate (R-3HB), is a fundamental constituent of biological cells. Certain prokaryotes accumulate PHB of very high molecular weight (10,000 to >1,000,000 residues), which is segregated within granular deposits in the cytoplasm; however, all prokaryotes and all eukaryotes synthesize PHB of medium-chain length (~100–200 residues) which resides within lipid bilayers or lipid vesicles, and PHB of short-chain length (<12 residues) which is conjugated to proteins (cPHB), primarily proteins in membranes and organelles. The physical properties of cPHB indicate it plays important roles in the targeting and folding of cPHB-proteins. Here we review the occurrence, physical properties and molecular characteristics of cPHB, and discuss its influence on the folding and structure of outer membrane protein A (OmpA) of Escherichia coli.

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Section: The Role Of Cphb In Folding Of Outer Membrane Protein a (mentioning

confidence: 68%

The Role of Short-Chain Conjugated Poly-(R)-3-Hydroxybutyrate (cPHB) in Protein Folding

Reusch

2013

IJMS

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“…A structural model of this domain has been developed and accepted by many researchers, which is based on a highly homologous protein RmpM from Neisseria meningitides (42). Despite these efforts, the oligomeric state of OmpA in vivo has remained unclear (32,40,(43)(44)(45). In general, membrane proteins are the most difficult proteins to study structurally, because they can be strongly influenced by the presence of the lipid bilayer, are difficult to overexpress and fold properly, and frequently contain disordered regions (46).…”

Section: Pir Results Of Protein Complexes Without Knownmentioning

confidence: 99%

Cross-linking Measurements of In Vivo Protein Complex Topologies

Zheng

Hoopmann

et al. 2011

Molecular & Cellular Proteomics

129

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Protein interactions and topologies are key features that enable specificity, function and the evolution of highly integrated, regulated networks in biological systems. Primary challenges associated with the study of biological systems include identification of protein interactions and measurement of topological features of proteins and their interactions in vivo. Advancements such as the Yeast Two-Hybrid (1), coimmunoprecipitation (2), and Tandem Affinity Purification tags (3) have greatly increased the ability to identify hundreds or even thousands of interactions from complex biological samples (2, 4 -6). Despite the many thousands of protein interactions that are now known (7) however, for only a tiny fraction is there any knowledge of their in vivo topology. On the other hand, if topologies of interactions were more widely known, this information could improve understanding of underlying fundamental factors that drive interactions, improve development of highly specific modulators of protein interactions, improve interaction prediction capabilities, and improve comprehension on biological systems. Unfortunately, exceedingly few methods exist to allow unbiased measurement of proteinprotein interaction topological features in cells.Chemical cross-linking has great potential for in vivo interaction topological studies (8 -10). Cross-linked peptides contain information about interacting protein identities and can uniquely define regions of protein sequences that are near one another when proteins are present within the native cellular environment. Challenges associated with in vivo crosslinking analysis that have precluded this achievement include the difficulty in identification of cross-linked peptides and the severe dynamic range constraints resultant from the overwhelming majority of noncross-linked peptides. Our efforts to overcome these challenges resulted in development of Protein Interaction Reporter (PIR) 1 technology (11) that uses a novel type of cross-linker and mass spectrometry to identify peptides that are close to one another within protein complexes in cells. These efforts resulted in the first reported identification of cross-linked peptides from live cells (9) including the first in vivo identification of an interaction among two outer membrane cytochrome c proteins, an interaction that appears to be critical to electron transport properties of Shewanella oneidensis (12).Here we present the first application of PIR technology to the study of interactions in E. coli cells where 65 cross-linked peptide pairs were unambiguously identified. To date, this constitutes the largest in vivo cross-linked peptide data set ever produced. In this system, we are also able to compare many of our results with known protein and protein complex crystal structures that demonstrate excellent agreement with our in vivo data. Importantly, this comparative analysis was also used to define distance constraints that enable refinements of structural prediction of in vivo protein complexes never before possible. Furthe...

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“…It is worth mentioning that the homologous OprF protein from Pseudomonas species has been shown to have a tendency to form an oligomer only when it forms a large pore [29]. To explain the appearance of the larger pore for E. coli OmpA, a model for a 16-stranded β-barrel structure that involves both the N-and C-portions of E. coli OmpA has been proposed [72,67]. However, such a transition would require the complete unfolding and refolding of the stable N-terminal β-barrel, as well as unfolding of the C-terminal domain, which is independently folded and also forms a stable structure.…”

Section: Discussionmentioning

confidence: 99%

The periplasmic domain of Escherichia coli outer membrane protein A can undergo a localized temperature dependent structural transition

Ishida

Garcı́a-Herrero

Vogel

2014

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Gram-negative bacteria such as Escherichia coli are surrounded by two membranes with a thin peptidoglycan (PG)-layer located in between them in the periplasmic space. The outer membrane protein A (OmpA) is a 325-residue protein and it is the major protein component of the outer membrane of E. coli. Previous structure determinations have focused on the N-terminal fragment (residues 1-171) of OmpA, which forms an eight stranded transmembrane β-barrel in the outer membrane. Consequently it was suggested that OmpA is composed of two independently folded domains in which the N-terminal β-barrel traverses the outer membrane and the C-terminal domain (residues 180-325) adopts a folded structure in the periplasmic space. However, some reports have proposed that full-length OmpA can instead refold in a temperature dependent manner into a single domain forming a larger transmembrane pore. Here, we have determined the NMR solution structure of the C-terminal periplasmic domain of E. coli OmpA (OmpA(180-325)). Our structure reveals that the C-terminal domain folds independently into a stable globular structure that is homologous to the previously reported PG-associated domain of Neisseria meningitides RmpM. Our results lend credence to the two domain structure model and a PG-binding function for OmpA, and we could indeed localize the PG-binding site on the protein through NMR chemical shift perturbation experiments. On the other hand, we found no evidence for binding of OmpA(180-325) with the TonB protein. In addition, we have also expressed and purified full-length OmpA (OmpA(1-325)) to study the structure of the full-length protein in micelles and nanodiscs by NMR spectroscopy. In both membrane mimetic environments, the recombinant OmpA maintains its two domain structure that is connected through a flexible linker. A series of temperature-dependent HSQC experiments and relaxation dispersion NMR experiments detected structural destabilization in the bulge region of the periplasmic domain of OmpA above physiological temperatures, which may induce dimerization and play a role in triggering the previously reported larger pore formation.

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An alternative topological model for Escherichia coli OmpA

Cited by 41 publications

References 39 publications

The Role of Short-Chain Conjugated Poly-(R)-3-Hydroxybutyrate (cPHB) in Protein Folding

The Role of Short-Chain Conjugated Poly-(R)-3-Hydroxybutyrate (cPHB) in Protein Folding

Cross-linking Measurements of In Vivo Protein Complex Topologies

The periplasmic domain of Escherichia coli outer membrane protein A can undergo a localized temperature dependent structural transition

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