scholarly journals Dynamic periplasmic chaperone reservoir facilitates biogenesis of outer membrane proteins

2016 ◽  
Vol 113 (33) ◽  
pp. E4794-E4800 ◽  
Author(s):  
Shawn M. Costello ◽  
Ashlee M. Plummer ◽  
Patrick J. Fleming ◽  
Karen G. Fleming
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Effective Rate ◽  
Stochastic Methods ◽  
Bacterial Physiology ◽  
Control Factors

Outer membrane protein (OMP) biogenesis is critical to bacterial physiology because the cellular envelope is vital to bacterial pathogenesis and antibiotic resistance. The process of OMP biogenesis has been studied in vivo, and each of its components has been studied in isolation in vitro. This work integrates parameters and observations from both in vivo and in vitro experiments into a holistic computational model termed “Outer Membrane Protein Biogenesis Model” (OMPBioM). We use OMPBioM to assess OMP biogenesis mathematically in a global manner. Using deterministic and stochastic methods, we are able to simulate OMP biogenesis under varying genetic conditions, each of which successfully replicates experimental observations. We observe that OMPs have a prolonged lifetime in the periplasm where an unfolded OMP makes, on average, hundreds of short-lived interactions with chaperones before folding into its native state. We find that some periplasmic chaperones function primarily as quality-control factors; this function complements the folding catalysis function of other chaperones. Additionally, the effective rate for the β-barrel assembly machinery complex necessary for physiological folding was found to be higher than has currently been observed in vitro. Overall, we find a finely tuned balance between thermodynamic and kinetic parameters maximizes OMP folding flux and minimizes aggregation and unnecessary degradation. In sum, OMPBioM provides a global view of OMP biogenesis that yields unique insights into this essential pathway.

scholarly journals Purification of the Major Outer Membrane Protein of Azospirillum brasilense, Its Affinity to Plant Roots, and Its Involvement in Cell Aggregation

2001 ◽  
Vol 14 (4) ◽  
pp. 555-561 ◽  
Author(s):  
Saul Burdman ◽  
Gabriella Dulguerova ◽  
Yaacov Okon ◽  
Edouard Jurkevitch
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Azospirillum Brasilense ◽  
Outer Membrane Proteins ◽  
Cell Aggregation ◽  
Major Outer Membrane Protein ◽  
Adhesion Assay ◽  
Fab Fragments

The major outer membrane protein (MOMP) of the nitrogen-fixing rhizobacterium Azospirillum brasilense strain Cd was purified and isolated by gel filtration, and antiserum against this protein was obtained. A screening of the binding of outer membrane proteins (OMPs) of A. brasilense to membrane-immobilized root extracts of various plant species revealed different affinities for the MOMP, with a stronger adhesion to extracts of cereals in comparison with legumes and tomatoes. Moreover, this protein was shown to bind to roots of different cereal seedlings in an in vitro adhesion assay. Incubation of A. brasilense cells with MOMP-antiserum led to fast agglutination, indicating that the MOMP is a surface-exposed protein. Cells incubated with Fab fragments obtained from purified MOMP-antiserum immunoglobulin G exhibited significant inhibition of bacterial aggregation as compared with controls. Bacteria preincubated with Fab fragments showed weaker adhesion to corn roots in comparison to controls without Fab fragments. These findings suggest that the A. brasilense MOMP acts as an adhesin involved in root adsorption and cell aggregation of this bacterium.

scholarly journals Characterization of Specific Immune Responses of Mice Inoculated with Recombinant Vaccinia Virus Expressing an 18-Kilodalton Outer Membrane Protein of Brucella abortus

2000 ◽  
Vol 7 (1) ◽  
pp. 114-118 ◽  
Author(s):  
Ramesh Vemulapalli ◽  
Silvio Cravero ◽  
Christine L. Calvert ◽  
Thomas E. Toth ◽  
Nammalwar Sriranganathan ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Vaccinia Virus ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Brucella Abortus ◽  
Virulent Strain ◽  
Shuttle Vector ◽  
Protein Fusion

ABSTRACT Using the shuttle vector pMCO2 and the vaccinia virus wild-type WR strain, we constructed a recombinant virus expressing an 18-kDa outer membrane protein of Brucella abortus. BALB/c mice inoculated with this virus produced 18-kDa protein-specific antibodies, mostly of immunoglobulin G2a isotype, and in vitro stimulation of splenocytes from these mice with purified maltose binding protein–18-kDa protein fusion resulted in lymphocyte proliferation and gamma interferon production. However, these mice were not protected against a challenge with the virulent strain B. abortus2308. Disruption of the 18-kDa protein's gene in vaccine strainB. abortus RB51 did not affect either the strain's protective capabilities or its in vivo attenuation characteristics. These observations suggest that the 18-kDa protein plays no role in protective immunity.

scholarly journals LipL32 Is an Extracellular Matrix-Interacting Protein of Leptospira spp. and Pseudoalteromonas tunicata

2008 ◽  
Vol 76 (5) ◽  
pp. 2063-2069 ◽  
Author(s):  
David E. Hoke ◽  
Suhelen Egan ◽  
Paul A. Cullen ◽  
Ben Adler
Keyword(s):  
Extracellular Matrix ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Major Outer Membrane Protein ◽  
Interacting Proteins ◽  
Orthologous Protein ◽  
Pseudoalteromonas Tunicata

ABSTRACT LipL32 is the major outer membrane protein in pathogenic Leptospira. It is highly conserved throughout pathogenic species and is expressed in vivo during human infection. While these data suggest a role in pathogenesis, a function for LipL32 has not been defined. Outer membrane proteins of gram-negative bacteria are the first line of molecular interaction with the host, and many have been shown to bind host extracellular matrix (ECM). A search for leptospiral ECM-interacting proteins identified the major outer membrane protein, LipL32. To verify this finding, recombinant LipL32 was expressed in Escherichia coli and was found to bind Matrigel ECM and individual components of ECM, including laminin, collagen I, and collagen V. Likewise, an orthologous protein found in the genome of Pseudoalteromonas tunicata strain D2 was expressed and found to be functionally similar and immunologically cross-reactive. Lastly, binding activity was mapped to the C-terminal 72 amino acids. These studies show that LipL32 and an orthologous protein in P. tunicata are immunologically cross-reactive and function as ECM-interacting proteins via a conserved C-terminal region.

scholarly journals Refolding of Escherichia coli outer membrane protein F in detergent creates LPS-free trimers and asymmetric dimers

2005 ◽  
Vol 392 (2) ◽  
pp. 375-381 ◽  
Author(s):  
Virak Visudtiphole ◽  
Matthew B. Thomas ◽  
David A. Chalton ◽  
Jeremy H. Lakey
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Single Molecule ◽  
Outer Membrane Protein ◽  
Structural Integrity ◽  
Protein F ◽  
Outer Membrane Protein F

The Escherichia coli OmpF (outer-membrane protein F; matrix porin) is a homotrimeric β-barrel and a member of the bacterial porin superfamily. It is the best characterized porin protein, but has resisted attempts to refold it efficiently in vitro. In the present paper, we report the discovery of detergent-based folding conditions, including dodecylglucoside, which can create pure samples of trimeric OmpF. Whereas outer membrane LPS (lipopolysaccharide) is clearly required for in vivo folding, the artificially refolded and LPS-free trimer has properties identical with those of the outer-membrane-derived form. Thus LPS is not required either for in vitro folding or for structural integrity. Dimeric forms of OmpF have been observed in vivo and are proposed to be folding intermediates. In vitro, dimers occur transiently in refolding of trimeric OmpF and, in the presence of dodecylmaltoside, pure dimer can be prepared. This form has less β-structure by CD and shows lower thermal stability than the trimer. Study of these proteins at the single-molecule level is possible because each OmpF subunit forms a distinct ion channel. Whereas each trimer contains three channels of equal conductance, each dimer always contains two distinct channel sizes. This provides clear evidence that the two otherwise identical monomers adopt different structures in the dimer and indicates that the asymmetric interaction, characteristic of C3 symmetry, is formed at the dimer stage. This asymmetric dimer may be generally relevant to the folding of oligomeric proteins with odd numbers of subunits such as aspartate transcarbamoylase.

scholarly journals Expression of Members of the 28-Kilodalton Major Outer Membrane Protein Family of Ehrlichia chaffeensis during Persistent Infection

2004 ◽  
Vol 72 (8) ◽  
pp. 4336-4343 ◽  
Author(s):  
Jian-zhi Zhang ◽  
Hong Guo ◽  
Gary M. Winslow ◽  
Xue-jie Yu
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Multigene Family ◽  
Outer Membrane Proteins ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ehrlichia Chaffeensis ◽  
Antigenic Peptides ◽  
Indirect Measure

ABSTRACT The 28-kDa immunodominant outer membrane proteins (P28 OMPs) of Ehrlichia chaffeensis are encoded by a multigene family. As an indirect measure of the in vivo expression of the members of the p28 multigene family of E. chaffeensis, sera from two beagle dogs experimentally infected with E. chaffeensis were evaluated for the presence of specific antibodies to P28 OMPs by enzyme-linked immunosorbent assay. Antigenic peptides unique to each of the P28s were identified within the first hypervariable region of each P28 OMP. Serological responses to peptides derived from all P28 OMPs were detected from day 30 postinoculation to day 468 and from day 46 until day 159 in the two beagles. Although antibody titers to the peptides fluctuated, the peak response to all of the peptides appeared simultaneously in each dog. The antibody responses to another outer membrane protein of E. chaffeensis (GP120) showed similar temporal and quantitative changes. These data suggest that the P28 OMPs are expressed concurrently during persistent Ehrlichia infection.

scholarly journals Screening of anti-Acinetobacter baumannii phytochemicals, based on the potential inhibitory effect on OmpA and OmpW functions

2021 ◽  
Author(s):  
Shahab Shahryari ◽  
Parvin Mohammadnejad ◽  
Kambiz Akbari Noghabi
Keyword(s):  
Membrane Protein ◽  
Acinetobacter Baumannii ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Simulation Analysis ◽  
Protein A ◽  
Binding Pocket ◽  
Dimensional Structure

Therapeutic options, including last-line or combined antibiotic therapies for multi-drug resistant (MDR) strains of Acinetobacter baumannii are ineffective. The outer membrane protein A (OmpA) and outer membrane protein W (OmpW) are two porins known for their different cellular functions. Identification of natural compounds with the potentials to block these putative porins can attenuate the growth of the bacteria and control the relating diseases. The current work aimed to screen a library of 384 phytochemicals according to their potentials to be used as a drug, and potentials to inhibit the function of OmpA and OmpW in A. baumannii. The phytocompounds were initially screened based on their physicochemical, absorption, distribution, metabolism, excretion, and toxicity (ADMET) drug-like properties. Afterward, the selected ligands were subjected to standard docking calculations against the predicted three-dimensional structure of OmpA and OmpW in A. baumannii. We identified three phytochemicals (isosakuranetin, aloe-emodin and pinocembrin) possessing appreciable binding affinity towards the selected binding pocket of OmpA and OmpW. Molecular dynamics (MD) simulation analysis confirmed the stability of the complexes. Amongst them, isosakuranetin was suggested as the best phytocompound for further in vitro and in vivo study.

scholarly journals Involvement of HxuC Outer Membrane Protein in Utilization of Hemoglobin by Haemophilus influenzae

2001 ◽  
Vol 69 (4) ◽  
pp. 2353-2363 ◽  
Author(s):  
Leslie D. Cope ◽  
Robert P. Love ◽  
Sarah E. Guinn ◽  
Andrei Gilep ◽  
Sergei Usanov ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Haemophilus Influenzae ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Open Reading Frames ◽  
Triple Mutant ◽  
Nthi Strain ◽  
Free Heme

ABSTRACT Haemophilus influenzae can utilize different protein-bound forms of heme for growth in vitro. A previous study from this laboratory indicated that nontypeable Haemophilus influenzae (NTHI) strain N182 expressed three outer membrane proteins, designated HgbA, HgbB, and HgbC, that bound hemoglobin or hemoglobin-haptoglobin and were encoded by open reading frames (ORFs) that contained a CCAA nucleotide repeat. Testing of mutants expressing the HgbA, HgbB, and HgbC proteins individually revealed that expression of any one of these proteins was sufficient to allow wild-type growth with hemoglobin. In contrast, mutants that expressed only HgbA or HgbC grew significantly better with hemoglobin-haptoglobin than did a mutant expressing only HgbB. Construction of an isogenic hgbA hgbB hgbC mutant revealed that the absence of these three gene products did not affect the ability of NTHI N182 to utilize hemoglobin as a source of heme, although this mutant was severely impaired in its ability to utilize hemoglobin-haptoglobin. The introduction of atonB mutation into this triple mutant eliminated its ability to utilize hemoglobin, indicating that the pathway for hemoglobin utilization in the absence of HgbA, HgbB, and HgbC involved a TonB-dependent process. Inactivation in this triple mutant of thehxuC gene, which encodes a predicted TonB-dependent outer membrane protein previously shown to be involved in the utilization of free heme, resulted in loss of the ability to utilize hemoglobin. The results of this study reinforce the redundant nature of the heme acquisition systems expressed by H. influenzae.

scholarly journals Screening of anti- Acinetobacter baumannii phytochemicals, based on the potential inhibitory effect on OmpA and OmpW functions

2021 ◽  
Vol 8 (8) ◽  
pp. 201652
Author(s):  
Shahab Shahryari ◽  
Parvin Mohammadnejad ◽  
Kambiz Akbari Noghabi
Keyword(s):  
Membrane Protein ◽  
Acinetobacter Baumannii ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Simulation Analysis ◽  
Protein A ◽  
Dynamics Simulation ◽  
Dimensional Structure

Therapeutic options including last-line or combined antibiotic therapies for multi-drug-resistant strains of Acinetobacter baumannii are ineffective. The outer membrane protein A (OmpA) and outer membrane protein W (OmpW) are two porins known for their different cellular functions. Identification of natural compounds with the potentials to block these putative porins can attenuate the growth of the bacteria and control the relating diseases. The current work aimed to screen a library of 384 phytochemicals according to their potentials to be used as a drug, and potentials to inhibit the function of OmpA and OmpW in A. baumannii . The phytocompounds were initially screened based on their physico-chemical, absorption, distribution, metabolism, excretion and toxicity (ADMET) drug-like properties. Afterwards, the selected ligands were subjected to standard docking calculations against the predicted three-dimensional structure of OmpA and OmpW in A. baumannii . We identified three phytochemicals (isosakuranetin, aloe-emodin and pinocembrin) possessing appreciable binding affinity towards the selected binding pocket of OmpA and OmpW. Molecular dynamics simulation analysis confirmed the stability of the complexes. Among them, isosakuranetin was suggested as the best phytocompound for further in vitro and in vivo study.

scholarly journals Involvement of a 43-kilodalton outer membrane protein in beta-lactam resistance of Shigella dysenteriae.

1997 ◽  
Vol 41 (10) ◽  
pp. 2302-2304 ◽  
Author(s):  
A K Kar ◽  
A S Ghosh ◽  
K Chauhan ◽  
J Ahamed ◽  
J Basu ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Shigella Dysenteriae ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Beta Lactam Antibiotics ◽  
Resistant Strains

A beta-lactam-sensitive strain (C152) of Shigella dysenteriae showed two major outer membrane proteins (OMPs) with M(r)s of 43,000 and 38,000, while the clinical isolate M2 lacked the 43,000-Mr OMP, which acted as a channel for beta-lactam antibiotics. Permeability of beta-lactams across the outer membrane (OM) of M2 was lower than that across the OM of C152. Mutants deficient in the 43-kDa OMP could be selected in vitro from strain C152 in the presence of cefoxitin. All beta-lactam-resistant strains were sensitive to imipenem.

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