scholarly journals The mating pair stabilization protein, TraN, of the F plasmid is an outer-membrane protein with two regions that are important for its function in conjugation

Microbiology ◽  
2005 ◽  
Vol 151 (11) ◽  
pp. 3527-3540 ◽  
Author(s):  
William A. Klimke ◽  
Candace D. Rypien ◽  
Barbara Klinger ◽  
R. Alexander Kennedy ◽  
J. Manuel Rodriguez-Maillard ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Recipient Cell ◽  
Terminal Region ◽  
Mating Pair ◽  
Secretion Systems ◽  
F Plasmid ◽  
Type Iv ◽  
Reducing Conditions

F plasmid TraN (602 aa, processed to 584 aa with 22 conserved cysteines), which is essential for F plasmid conjugation, is an outer-membrane protein involved in mating pair stabilization (MPS). Unlike R100 TraN, F TraN requires OmpA in the recipient cell for efficient MPS. The authors have identified three external loops (aa 172–187, 212–220 and 281–284) in the highly divergent region from aa 164 to aa 333 as candidates for interaction with OmpA. These loops were identified using both site-directed and random TnphoA/in mutagenesis to insert epitopes (31-aa or c-myc) into TraN and monitor their effect on sensitivity to external proteases and on mating ability. TraN is a hallmark protein of F-type IV secretion systems as demonstrated by blast searches of the databases. The C-terminal region is highly conserved and contains five of the six completely conserved cysteines. Mutation of these residues to serine demonstrated their importance in TraN function. TraN appears to require both intra- and intermolecular disulfide bond formation for its stability and structure as demonstrated by its instability in a dsbA mutant and its aberrant migration on SDS-polyacrylamide gels under non-reducing conditions or by cross-linking with bis(sulfosuccinimidyl)suberate (BS3). Thus, F TraN appears to have two domains: the N-terminal region is involved in OmpA interaction with OmpA during MPS; and the C-terminal region, which is rich in conserved cysteine residues, is essential for conjugation.

Quantification and surface localization of the hemolysin A type 1 secretion system at the endogenous level and under conditions of overexpression

2021 ◽  
Author(s):  
Tobias Beer ◽  
Sebastian Hänsch ◽  
Klaus Pfeffer ◽  
Sander H.J. Smits ◽  
Stefanie Weidtkamp-Peters ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Extracellular Space ◽  
Super Resolution ◽  
Secretion Systems ◽  
Endogenous Level ◽  
Surface Localization ◽  
Bacterial Secretion

Secretion systems are essential for Gram-negative bacteria as these nanomachineries allow a communication with the outside world by exporting proteins into the extracellular space or directly into the cytosol of a host cell. For example, type one secretion systems (T1SS) secrete a broad range of substrates across both membranes into the extracellular space. One well-known example is the hemolysin A (HlyA) T1SS from Escherichia coli (E. coli) , which consists of an ABC transporter (HlyB), a membrane fusion protein (HlyD), the outer membrane protein TolC and the substrate HlyA, a member of the family of RTX (repeats in toxins) toxins. Here, we determined the amount of TolC at the endogenous level (parental strain, UTI89) and under conditions of overexpression (T7 expression system, BL21(DE3)-BD). The overall amount of TolC was not influenced by the overexpression of the HlyBD complex. Moving one step further, we determined the localization of the HlyA T1SS by super-resolution microscopy. In contrast to other bacterial secretion systems, no polarization was observed with respect to endogenous or overexpression levels. Additionally, the cell growth and division cycle did not influence the polarization. Most importantly, the size of the observed T1SS clusters did not correlate with the recently proposed outer membrane islands. These data indicate that T1SS cluster at the outer membrane generating domains of so far not described identity. Importance Uropathogenic Escherichia coli (UPEC) strains cause about 110 million urinary tract infections each year worldwide representing a global burden to the healthcare system. UPEC secrete many virulence factors among these the TX toxin hemolysin A via a cognate T1SS into the extracellular space. In this study, we determined the endogenous copy number of the HlyA T1SS in UTI89 and analyzed the surface localization in BL21(DE3)-BD and UTI89, respectively. With approximately 800 copies of the T1SS in UTI89, this is one of the highest expressed bacterial secretion systems. Furthermore and in clear contrast to other secretion systems, no polarized surface localization was detected. Finally, quantitative analysis of the super-resolution data revealed that clusters of the HlyA T1SS are not related to the recently identified outer membrane protein islands. These data provide insights into the quantitative molecular architecture of the HlyA T1SS.

scholarly journals Role of the carboxy-terminal region of the outer membrane protein AatA in the export of dispersin from enteroaggregative Escherichia coli

2006 ◽  
Vol 256 (2) ◽  
pp. 266-272 ◽  
Author(s):  
Mayumi Iwashita ◽  
Junichiro Nishi ◽  
Naoko Wakimoto ◽  
Rika Fujiyama ◽  
Kimie Yamamoto ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Terminal Region ◽  
Carboxy Terminal Region ◽  
Carboxy Terminal

scholarly journals Toxin secretion and trafficking by Mycobacterium tuberculosis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
David Pajuelo ◽  
Uday Tak ◽  
Lei Zhang ◽  
Olga Danilchanka ◽  
Anna D. Tischler ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Molecular Mechanisms ◽  
Comprehensive Analysis ◽  
Secretion Systems ◽  
Type Vii Secretion ◽  
Toxin Secretion ◽  
Phagosomal Membrane

AbstractThe tuberculosis necrotizing toxin (TNT) is the major cytotoxicity factor of Mycobacterium tuberculosis (Mtb) in macrophages. TNT is the C-terminal domain of the outer membrane protein CpnT and gains access to the cytosol to kill macrophages infected with Mtb. However, molecular mechanisms of TNT secretion and trafficking are largely unknown. A comprehensive analysis of the five type VII secretion systems of Mtb revealed that the ESX-4 system is required for export of CpnT and surface accessibility of TNT. Furthermore, the ESX-2 and ESX-4 systems are required for permeabilization of the phagosomal membrane in addition to the ESX-1 system. Thus, these three ESX systems need to act in concert to enable trafficking of TNT into the cytosol of Mtb-infected macrophages. These discoveries establish new molecular roles for the two previously uncharacterized type VII secretion systems ESX-2 and ESX-4 and reveal an intricate link between toxin secretion and phagosomal permeabilization by Mtb.

scholarly journals Characterization of a Predominant Immunogenic Outer Membrane Protein of Riemerella anatipestifer

2000 ◽  
Vol 7 (2) ◽  
pp. 168-174 ◽  
Author(s):  
Sumathi Subramaniam ◽  
Bin Huang ◽  
Hilda Loh ◽  
Jimmy Kwang ◽  
Hai-Meng Tan ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Calcium Binding ◽  
Outer Membrane Proteins ◽  
Terminal Region ◽  
Gene Encoding ◽  
Riemerella Anatipestifer ◽  
Binding Domains

ABSTRACT The ompA gene, encoding the 42-kDa major antigenic outer membrane protein OmpA of Riemerella anatipestifer, the etiololgical agent of septicemia anserum exsudativa, was cloned and expressed in Escherichia coli. Recombinant OmpA displayed a molecular mass similar to that predicted from the nucleotide sequence of the ompA gene but lower than that observed in total cell lysates of R. anatipestifer. The ompA gene showed a conserved C-terminal region comprising the OmpA-like domain and a variable N-terminal region. This structure is similar to those of the analogous outer membrane proteins of several gram-negative bacteria. However, OmpA of R. anatipestifercontains six EF-hand calcium-binding domains and two PEST regions, which distinguish it from other outer membrane proteins. The occurrence of these motifs in OmpA suggests a possible role in virulence for this protein. The ompA gene is present in the R. anatipestifer type strain and in all serotype reference strains. However, it exhibits some minor genetic heterogeneity among different serotypes, which seems not to affect the strong antigenic characteristics of the protein. OmpA is a conserved and strong antigenic determinant of R. anatipestifer and hence is suggested to be a valuable protein for the serodetection of R. anatipestifer infections, independent of their serotype.

Molecular insights into type I secretion systems

2013 ◽  
Vol 394 (11) ◽  
pp. 1371-1384 ◽  
Author(s):  
Michael H.H. Lenders ◽  
Sven Reimann ◽  
Sander H. J. Smits ◽  
Lutz Schmitt
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Atp Binding ◽  
Type I ◽  
Secretion Systems ◽  
Inner Membrane ◽  
Small Proteins ◽  
Atp Binding Cassette Transporter ◽  
Atp Binding Cassette

Abstract Type 1 secretion systems are one of the main machineries in Gram-negative bacteria involved in the secretion of a wide range of substrates from the cytoplasm across the inner and outer membrane in one step to the extracellular space. The range of substrates varies from small proteins up to large surface layer proteins of about 900 kDa. Most of the substrates have a non-cleavable C-terminal secretion signal and so-called GG repeats that are able to bind calcium ions. The translocator complex is composed of a trimeric outer membrane protein that provides a pore in the outer membrane. A multimeric membrane fusion protein spans the periplasm and forms a continuous channel connecting the outer membrane protein with a dimeric ATP-binding cassette transporter in the inner membrane. The ATP-binding cassette-transporter is thought to form a channel through the inner membrane and energizes the transport process. This review will provide a detailed view of the components of the translocator and will summarize structural as well as functional data.

P-217 Role of Adhesin of Outer Membrane Protein Chlamydia pneumoniae in Collagen Type-IV Degradation

2009 ◽  
Vol 4 ◽  
pp. S113
Author(s):  
Sri Murwani ◽  
Djanggan Sargowo ◽  
Handono Kalim ◽  
Mulyohadi Ali ◽  
Ketut Muliartha ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Chlamydia Pneumoniae ◽  
Collagen Type ◽  
Collagen Type Iv ◽  
Type Iv

scholarly journals 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

Microbiology ◽  
2010 ◽  
Vol 156 (3) ◽  
pp. 742-756 ◽  
Author(s):  
Kathleen R. Ryan ◽  
James A. Taylor ◽  
Lisa M. Bowers
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Caulobacter Crescentus ◽  
Heat Exposure ◽  
Membrane Integrity ◽  
Permeability Barrier ◽  
Type Iv ◽  
Bam Complex ◽  
Selective Permeability

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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