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 Sequence Polymorphism, Predicted Secondary Structures, and Surface-Exposed Conformational Epitopes of Campylobacter Major Outer Membrane Protein

2000 ◽  
Vol 68 (10) ◽  
pp. 5679-5689 ◽  
Author(s):  
Qijing Zhang ◽  
Jerrel C. Meitzler ◽  
Shouxiong Huang ◽  
Teresa Morishita
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Major Outer Membrane Protein ◽  
Amino Acid Sequences ◽  
Variable Regions ◽  
Conserved Sequences ◽  
Conformational Epitopes

ABSTRACT The major outer membrane protein (MOMP), a putative porin and a multifunction surface protein of Campylobacter jejuni, may play an important role in the adaptation of the organism to various host environments. To begin to dissect the biological functions and antigenic features of this protein, the gene (designatedcmp) encoding MOMP was identified and characterized from 22 strains of C. jejuni and one strain of C. coli. It was shown that the single-copy cmp locus encoded a protein with characteristics of bacterial outer membrane proteins. Prediction from deduced amino acid sequences suggested that each MOMP subunit consisted of 18 β-strands connected by short periplasmic turns and long irregular external loops. Alignment of the amino acid sequences of MOMP from different strains indicated that there were seven localized variable regions dispersed among highly conserved sequences. The variable regions were located in the putative external loop structures, while the predicted β-strands were formed by conserved sequences. The sequence homology of cmp appeared to reflect the phylogenetic proximity of C. jejuni strains, since strains with identical cmp sequences had indistinguishable or closely related macrorestriction fragment patterns. Using recombinant MOMP and antibodies recognizing linear or conformational epitopes of the protein, it was demonstrated that the surface-exposed epitopes of MOMP were predominantly conformational in nature. These findings are instrumental in the design of MOMP-based diagnostic tools and vaccines.

scholarly journals A novel O -linked glycan modulates Campylobacter jejuni major outer membrane protein-mediated adhesion to human histo-blood group antigens and chicken colonization

Open Biology ◽  
2014 ◽  
Vol 4 (1) ◽  
pp. 130202 ◽  
Author(s):  
Jafar Mahdavi ◽  
Necmettin Pirinccioglu ◽  
Neil J. Oldfield ◽  
Elisabet Carlsohn ◽  
Jeroen Stoof ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Blood Group ◽  
Campylobacter Jejuni ◽  
Outer Membrane Protein ◽  
Ex Vivo ◽  
Binding Capacity ◽  
Major Outer Membrane Protein ◽  
Blood Group Antigens

Campylobacter jejuni is an important cause of human foodborne gastroenteritis; strategies to prevent infection are hampered by a poor understanding of the complex interactions between host and pathogen. Previous work showed that C. jejuni could bind human histo-blood group antigens (BgAgs) in vitro and that BgAgs could inhibit the binding of C. jejuni to human intestinal mucosa ex vivo. Here, the major flagella subunit protein (FlaA) and the major outer membrane protein (MOMP) were identified as BgAg-binding adhesins in C. jejuni NCTC11168 . Significantly, the MOMP was shown to be O- glycosylated at Thr 268 ; previously only flagellin proteins were known to be O- glycosylated in C. jejuni . Substitution of MOMP Thr 268 led to significantly reduced binding to BgAgs. The O- glycan moiety was characterized as Gal(β1–3)-GalNAc(β1–4)-GalNAc(β1–4)-GalNAcα1-Thr 268 ; modelling suggested that O- glycosylation has a notable effect on the conformation of MOMP and this modulates BgAg-binding capacity. Glycosylation of MOMP at Thr 268 promoted cell-to-cell binding, biofilm formation and adhesion to Caco-2 cells, and was required for the optimal colonization of chickens by C. jejuni , confirming the significance of this O- glycosylation in pathogenesis.

scholarly journals Characterization of Outer Membrane Proteins in Chlamydia trachomatis LGV Serovar L2

2001 ◽  
Vol 183 (8) ◽  
pp. 2686-2690 ◽  
Author(s):  
Regina J. Tanzer ◽  
Thomas P. Hatch
Keyword(s):  
Chlamydia Trachomatis ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Major Outer Membrane Protein ◽  
Developmental Cycle ◽  
Reading Frame

ABSTRACT We used a photoactivatable, lipophilic reagent, 3′-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine, to label proteins in the outer membrane of elementary bodies ofChlamydia trachomatis LGV serovar L2 and mass spectrometry to identify the labeled proteins. The identified proteins were polymorphic outer membrane proteins E, G, and H, which were made late in the developmental cycle, the major outer membrane protein, and a mixture of 46-kDa proteins consisting of the open reading frame 623 protein and possibly a modified form of the major outer membrane protein.

scholarly journals Mutation of the gene encoding a major outer-membrane protein in Xanthomonas campestris pv. campestris causes pleiotropic effects, including loss of pathogenicity

Microbiology ◽  
2010 ◽  
Vol 156 (9) ◽  
pp. 2842-2854 ◽  
Author(s):  
Yih-Yuan Chen ◽  
Chieh-Hao Wu ◽  
Juey-Wen Lin ◽  
Shu-Fen Weng ◽  
Yi-Hsiung Tseng
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Xanthomonas Campestris ◽  
Extracellular Enzymes ◽  
Surface Deformation ◽  
Cell Signalling ◽  
Cell Aggregation ◽  
Major Outer Membrane Protein ◽  
Gene Encoding

Xanthomonas campestris pv. campestris (Xcc) is the phytopathogen that causes black rot in crucifers. The xanthan polysaccharide and extracellular enzymes produced by this organism are virulence factors, the expression of which is upregulated by Clp (CRP-like protein) and DSF (diffusible signal factor), which is synthesized by RpfF. It is also known that biofilm formation/dispersal, regulated by the effect of controlled synthesis of DSF on cell–cell signalling, is required for virulence. Furthermore, a deficiency in DSF causes cell aggregation with concomitant production of a gum-like substance that can be dispersed by addition of DSF or digested by exogenous endo-β-1,4-mannanase expressed by Xcc. In this study, Western blotting of proteins from a mopB mutant (XcMopB) showed Xcc MopB to be the major outer-membrane protein (OMP); Xcc MopB shared over 97 % identity with homologues from other members of Xanthomonas. Similarly to the rpfF mutant, XcMopB formed aggregates with simultaneous production of a gummy substance, but these aggregates could not be dispersed by DSF or endo-β-1,4-mannanase, indicating that different mechanisms were involved in aggregation. In addition, XcMopB showed surface deformation, altered OMP composition, impaired xanthan production, increased sensitivity to stressful conditions including SDS, elevated temperature and changes in pH, reduced adhesion and motility and defects in pathogenesis. The finding that the major OMP is required for pathogenicity is unprecedented in phytopathogenic bacteria.

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 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 Protection of Wild-Type and Severe Combined Immunodeficiency Mice against an Intranasal Challenge by Passive Immunization with Monoclonal Antibodies to the Chlamydia trachomatis Mouse Pneumonitis Major Outer Membrane Protein

2008 ◽  
Vol 76 (12) ◽  
pp. 5581-5587 ◽  
Author(s):  
Sukumar Pal ◽  
Jose Bravo ◽  
Ellena M. Peterson ◽  
Luis M. de la Maza
Keyword(s):  
Body Weight ◽  
Monoclonal Antibodies ◽  
Chlamydia Trachomatis ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Passive Immunization ◽  
Major Outer Membrane Protein ◽  
Wild Type

ABSTRACT Monoclonal antibodies (MAbs) to the Chlamydia trachomatis mouse pneumonitis (MoPn) major outer membrane protein (MOMP) were characterized for their ability to neutralize the infectivity of this organism in vitro and in vivo. One of the MAbs (MoPn-23) recognizes a nonlinear epitope in the MOMP, MAb MoPn-40 binds to a linear epitope in the variable domain 1 (VD1), and MAb MoPn-32 recognizes the chlamydial lipopolysaccharide. MAb MoPn-23 neutralized 50% of the infectivity of Chlamydia, as measured in vitro by using HAK (FcγIII−) and HeLa-229 (FcγIII+) cells at a concentration 100 times lower than MAb MoPn-40. MAb MoPn-32 had no neutralizing ability. In comparison to the control normal mouse immunoglobulin G, passive immunization of BALB/c mice with MAb MoPn-23 resulted in a highly significant protection against an intranasal (i.n.) challenge as determined by the change in body weight, the weight of the lungs, and the yield of Chlamydia inclusion-forming units (IFU) from the lungs. Passive immunization with MAb MoPn-40 resulted in a lower degree of protection, and MAb MoPn-32 afforded no protection. MAb MoPn-23 was also tested for its ability to protect wild-type (WT) and severe combined immunodeficient (SCID) C.B-17 mice against an i.n. challenge. Protection based on total body weight, lung weight, and yield of Chlamydia IFU was as effective in SCID as in WT C.B-17 mice. In conclusion, antibodies to MOMP can protect mice against a chlamydial infection in the presence or absence of T and B cells.

scholarly journals Functional refolding of the Campylobacter jejuni MOMP (major outer membrane protein) porin by GroEL from the same species

2004 ◽  
Vol 378 (3) ◽  
pp. 851-856 ◽  
Author(s):  
Florence GOULHEN ◽  
Emmanuelle DÉ ◽  
Jean-Marie PAGÈS ◽  
Jean-Michel BOLLA
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Lipid Bilayers ◽  
Campylobacter Jejuni ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Major Outer Membrane Protein ◽  
Molar Ratio ◽  
Maximum Efficiency ◽  
Transmission Electron

Functional and structural studies of outer membrane proteins from Gram-negative bacteria are frequently carried out using refolded proteins. Recombinant proteins are produced in Escherichia coli as inclusion bodies and then tediously refolded by dilution in buffered detergent solutions. In the present work, we obtained the refolding of MOMP (major outer membrane protein) from Campylobacter assisted by the molecular chaperone GroEL. Refolded MOMP recovered its native pore-forming activity when reconstituted in planar lipid bilayers. Both proteins were purified from the Campylobacter jejuni strain 85H. The purity of GroEL was assessed by silver staining and MS. Its native ultrastructure was observed by the use of transmission electron microscopy. Denaturation of MOMP was performed in urea at 65 °C followed by dialysis against 100 mM acetic acid, and was assessed by CD analysis. MOMP refolding reached a maximum efficiency in the presence of GroEL (at a MOMP/GroEL molar ratio of 9:1) and ATP. Under these conditions, 95% of denatured MOMP was refolded after a 15 min incubation. This approach represents an alternative method in studies of membrane protein refolding.

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