scholarly journals Phase Variation of PorA, a Major Outer Membrane Protein, Mediates Escape of Bactericidal Antibodies by Neisseria meningitidis

2013 ◽  
Vol 81 (4) ◽  
pp. 1374-1380 ◽  
Author(s):  
Isfahan Tauseef ◽  
Youssif M. Ali ◽  
Christopher D. Bayliss
Keyword(s):  
Membrane Protein ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Phase Variation ◽  
Surface Expression ◽  
Content Type ◽  
Repeat Tract ◽  
Fold Reduction

ABSTRACTSeveral outer membrane proteins ofNeisseria meningitidisare subject to phase variation due to alterations in simple sequence repeat tracts. The PorA protein is a major outer membrane protein and a target for protective host immune responses. Phase variation of PorA is mediated by a poly-G repeat tract present within the promoter, leading to alterations in protein expression levels.N. meningitidisstrain 8047 was subjected to serial passage in the presence of P1.2, a PorA-specific bactericidal monoclonal antibody. Rapid development of resistance to bactericidal activity was associated with a switch in the PorA repeat tract from 11G to 10G. Phase variants with a 10G repeat tract exhibited a 2-fold reduction in surface expression of PorA protein. AmutSmutant of strain 8047, with an elevated phase variation rate, exhibited a higher rate of escape and an association of escape with 10G and 9G variants, the latter having a 13-fold reduction in surface expression of PorA. We conclude that graduated reductions in the surface expression of outer membrane proteins mediated by phase variation enable meningococci to escape killingin vitroby bactericidal antibodies. These findings indicate how phase variation could have a major impact on immune escape and host persistence of meningococci.

scholarly journals An Extracytoplasmic Function Sigma Factor Controls Cellulose Utilization by Regulating the Expression of an Outer Membrane Protein inCytophaga hutchinsonii

2018 ◽  
Vol 85 (5) ◽  
Author(s):  
Xia Wang ◽  
Weixin Zhang ◽  
Hong Zhou ◽  
Guanjun Chen ◽  
Weifeng Liu
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Regulatory Mechanism ◽  
Outer Membrane Proteins ◽  
Cellulolytic Bacterium ◽  
Content Type ◽  
Cytophaga Hutchinsonii ◽  
Σ Factor ◽  
The Common

ABSTRACTThe common soil cellulolytic bacterium known asCytophaga hutchinsoniimakes use of a unique but poorly understood strategy in order to utilize cellulose. While several genes have been identified as being an active part of the utilization of cellulose, the mechanism(s) by whichC. hutchinsoniiboth (i) senses its environment and (ii) regulates the expression of those genes are not as yet known. In this study, we identified and characterized the geneCHU_3097encoding an extracytoplasmic function (ECF) σ factor (σcel1), the disruption of which compromisedC. hutchinsoniicellulose assimilation to a large degree. The σcel1and its putative partner anti-σcel1, encoded by theCHU_3096gene found immediately downstream fromCHU_3097, copurifiedin vitro. The σcel1was discovered to be associated with inner membrane when cells were cultured on glucose and yet was partially released from the membrane in response to cellulose. This release was found to occur on glucose when the anti-σcel1was absent. Transcriptome analyses found a σcel1-regulated, cellulose-responsive gene regulon, within which an outer membrane protein encoding the geneCHU_1276, essential for cellulose utilization, was discovered to be significantly downregulated byCHU_3097disruption. The expression of CHU_1276 almost fully restored cellulose utilization to theCHU_3097mutant, demonstrating that CHU_1276 represents a critical regulatory target of σcel1. In this way, our study provided insights into the role of an ECF σ factor in coordinating the cellulolytic response ofC. hutchinsonii.IMPORTANCEThe common cellulolytic bacteriumCytophaga hutchinsoniiuses a unique but poorly understood strategy in order to make use of cellulose. Throughout the process of cellulosic biomass breakdown, outer membrane proteins are thought to play key roles; this is evidenced by CHU_1276, which is required for the utilization of cellulose. However, the regulatory mechanism of its expression is not yet known. We found and characterized an extracytoplasmic function σ factor that is involved in coordinating the cellulolytic response ofC. hutchinsoniiby directly regulating the expression ofCHU_1276. This study makes a contribution to our understanding of the regulatory mechanism used byC. hutchinsoniiin order to adjust its genetic programs and so deal with novel environmental cues.

scholarly journals HmbR, a Hemoglobin-Binding Outer Membrane Protein of Neisseria meningitidis, Undergoes Phase Variation

1999 ◽  
Vol 181 (7) ◽  
pp. 2067-2074 ◽  
Author(s):  
Anthony R. Richardson ◽  
Igor Stojiljkovic
Keyword(s):  
Membrane Protein ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Phase Variation ◽  
Membrane Receptors ◽  
Phenotypic Switch ◽  
Reading Frame ◽  
Single Colony ◽  
Hemoglobin Binding

ABSTRACT Neisseria meningitidis uses hemoglobin (Hb) as an iron source via two TonB-dependent outer membrane receptors, HmbR and HpuB. Analysis of 25 epidemiologically unrelated clinical isolates from serogroups A, B, C, and Y revealed that 64% strains possessed both Hb receptor genes. Examination of the hmbR expression pattern in strains in which the hpuB gene was genetically inactivated revealed two distinct Hb utilization phenotypes. Five strains retained the ability to grow as a confluent lawn, while seven grew only as single colonies around Hb discs. The single-colony phenotype observed for some hpuB mutants is suggestive of phase variation of hmbR. The length of the poly(G) tract starting at position +1164 of hmbR absolutely correlated with the two Hb utilization phenotypes. All five strains that grew as confluent lawns around Hb discs possessed either 9 or 12 consecutive G residues. All seven strains that grew as single colonies around Hb discs had poly(G) tracts of a length other than 9 or 12. These single-colony variants that arose around the Hb discs had poly(G) tracts with either 9 or 12 consecutive G residues restoring thehmbR reading frame. Inactivation of hmbR in these strains resulted in a loss of Hb utilization, demonstrating that the change in the hmbR gene was responsible for the phenotypic switch. The switching rates from hmbR phase off to phase on were ∼5 × 10−4 in four serogroup C strains, 2 × 10−2 in the serogroup A isolate, and 7 × 10−6 in the serogroup B isolate.

scholarly journals Novel Outer Membrane Protein Involved in Cellulose and Cellooligosaccharide Degradation by Cytophaga hutchinsonii

2014 ◽  
Vol 80 (15) ◽  
pp. 4511-4518 ◽  
Author(s):  
Xiaofei Ji ◽  
Ying Wang ◽  
Cong Zhang ◽  
Xinfeng Bai ◽  
Weican Zhang ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Cell Surface ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Cellulose Degradation ◽  
Outer Membrane Proteins ◽  
Cellulolytic Enzymes ◽  
Content Type ◽  
Cytophaga Hutchinsonii ◽  
Glucosidase Activity

ABSTRACTCytophaga hutchinsoniiis an aerobic cellulolytic soil bacterium which was reported to use a novel contact-dependent strategy to degrade cellulose. It was speculated that cellooligosaccharides were transported into the periplasm for further digestion. In this study, we reported that most of the endoglucanase and β-glucosidase activity was distributed on the cell surface ofC. hutchinsonii. Cellobiose and part of the cellulose could be hydrolyzed to glucose on the cell surface. However, the cell surface cellulolytic enzymes were not sufficient for cellulose degradation byC. hutchinsonii. An outer membrane protein, CHU_1277, was disrupted by insertional mutation. Although the mutant maintained the same endoglucanase activity and most of the β-glucosidase activity, it failed to digest cellulose, and its cellooligosaccharide utilization ability was significantly reduced, suggesting that CHU_1277 was essential for cellulose degradation and played an important role in cellooligosaccharide utilization. Further study of cellobiose hydrolytic ability of the mutant on the enzymatic level showed that the β-glucosidase activity in the outer membrane of the mutant was not changed. It revealed that CHU_1277 played an important role in assisting cell surface β-glucosidase to exhibit its activity sufficiently. Studies on the outer membrane proteins involved in cellulose and cellooligosaccharide utilization could shed light on the mechanism of cellulose degradation byC. hutchinsonii.

scholarly journals Type-specific antigens of group A Neisseria meningitidis: lipopolysaccharide and heat-modifiable outer membrane proteins

1980 ◽  
Vol 28 (2) ◽  
pp. 451-458 ◽  
Author(s):  
W D Zollinger ◽  
R E Mandrell
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Solid Phase ◽  
Outer Membrane Proteins ◽  
Surface Antigens ◽  
Protein Antigens ◽  
Group A

The solid-phase radioimmunoassay inhibition method was used to analyze the noncapsular surface antigens of group A Neisseria meningitidis for type specificity. By use of antisera prepared against group A strains, three serologically distinct lipopolysaccharide antigens and five outer membrane protein antigens were identified among group A strains from a variety of geographical origins. Two of the lipopolysaccharide antigens were unique to group A strains while the third was similar to those on strains of other meningococcal serogroups. Fractionation of outer membrane proteins in the presence of 2% sodium deoxycholate followed by quantitative inhibition of the typing reactions with the subfractions revealed that the protein responsible for type specificity was not the principal outer membrane protein, but, most likely, the 31,000-dalton, heat-modifiable outer membrane protein. Thus, although group A strains may share a common principal outer membrane protein, typing is feasible using other surface antigens. In a survey of 82 group A strains, 93% were typable with respect to outer membrane proteins.

scholarly journals Evaluation of Protective Potential of Yersinia pestis Outer Membrane Protein Antigens as Possible Candidates for a New-Generation Recombinant Plague Vaccine

2012 ◽  
Vol 20 (2) ◽  
pp. 227-238 ◽  
Author(s):  
Tatiana E. Erova ◽  
Jason A. Rosenzweig ◽  
Jian Sha ◽  
Giovanni Suarez ◽  
Johanna C. Sierra ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Yersinia Pestis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Spectrometric Analysis ◽  
Protein Antigens ◽  
Pneumonic Plague ◽  
Content Type ◽  
New Generation

ABSTRACTPlague caused byYersinia pestismanifests itself in bubonic, septicemic, and pneumonic forms. Although the U.S. Food and Drug Administration recently approved levofloxacin, there is no approved human vaccine against plague. The capsular antigen F1 and the low-calcium-response V antigen (LcrV) ofY. pestisrepresent excellent vaccine candidates; however, the inability of the immune responses to F1 and LcrV to provide protection againstY. pestisF1−strains or those which harbor variants of LcrV is a significant concern. Here, we show that the passive transfer of hyperimmune sera from rats infected with the plague bacterium and rescued by levofloxacin protected naive animals against pneumonic plague. Furthermore, 10 to 12 protein bands from wild-type (WT)Y. pestisCO92 reacted with the aforementioned hyperimmune sera upon Western blot analysis. Based on mass spectrometric analysis, four of these proteins were identified as attachment invasion locus (Ail/OmpX), plasminogen-activating protease (Pla), outer membrane protein A (OmpA), and F1. The genes encoding these proteins were cloned, and the recombinant proteins purified fromEscherichia colifor immunization purposes before challenging mice and rats with either the F1−mutant or WT CO92 in bubonic and pneumonic plague models. Although antibodies to Ail and OmpA protected mice against bubonic plague when challenged with the F1−CO92 strain, Pla antibodies were protective against pneumonic plague. In the rat model, antibodies to Ail provided protection only against pneumonic plague after WT CO92 challenge. Together, the addition ofY. pestisouter membrane proteins to a new-generation recombinant vaccine could provide protection against a wide variety ofY. pestisstrains.

Sequencing ofporAfrom clinical isolates ofNeisseria meningitidisdefines a subtyping scheme and its genetic regulation

1998 ◽  
Vol 44 (1) ◽  
pp. 56-63 ◽  
Author(s):  
Francis F Arhin ◽  
France Moreau ◽  
James W Coulton ◽  
Elaine L Mills
Keyword(s):  
Membrane Protein ◽  
Dna Sequencing ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Promoter Region ◽  
Outer Membrane Proteins ◽  
Coding Region ◽  
Variable Regions ◽  
Class 1

Subtyping Neisseria meningitidis by methods that rely on monoclonal antibody (mAb) reactivity results in an unusually high number of strains that are not subtypeable. To subtype 48 strains isolated (1993-1994) in the province of Quebec that were not subtypeable by mAb-based techniques, we used DNA sequencing of the variable regions of porA, a gene that encodes the class 1 outer membrane protein. We assigned subtypes to all the previously nonserosubtypeable isolates and identified some novel subtypes. Because our sequencing strategy included the promoter region of porA, different isolates were compared in their sequences of the porA promoter region. A poly(G) stretch lies between the -10 and -35 regions of the promoter; replacement of a G residue by an A residue in this region resulted in loss of expression of porA. No correlation was found between the number of G residues in the poly(G) stretch and the level of expression; a minimum of 10 G residues is required in this stretch for expression of porA. One isolate expressed no class 1 outer membrane protein because of the insertion sequence IS1301 in the coding region of porA. Another isolate did not express the protein owing to a frame-shift mutation within the coding region of porA. Sequencing of porA allowed assignments of subtypes to previously uncharacterized isolates and provided insights about the regulation of expression of this gene in N. meningitidis.Key words: Neisseria meningitidis, outer membrane proteins, subtyping, PorA, DNA sequencing.

scholarly journals Two Protein Lysine Methyltransferases Methylate Outer Membrane Protein B from Rickettsia

2012 ◽  
Vol 194 (23) ◽  
pp. 6410-6418 ◽  
Author(s):  
Amila H. Abeykoon ◽  
Chien-Chung Chao ◽  
Guanghui Wang ◽  
Marjan Gucek ◽  
David C. H. Yang ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Dna Sequences ◽  
Outer Membrane Protein ◽  
Specific Activity ◽  
Outer Membrane Proteins ◽  
Enzymatic System ◽  
Content Type ◽  
Lysine Methyltransferases ◽  
Protein B

ABSTRACTRickettsia prowazekii, the etiologic agent of epidemic typhus, is a potential biological threat agent. Its outer membrane protein B (OmpB) is an immunodominant antigen and plays roles as protective envelope and as adhesins. The observation of the correlation between methylation of lysine residues in rickettsial OmpB and bacterial virulence has suggested the importance of an enzymatic system for the methylation of OmpB. However, no rickettsial lysine methyltransferase has been characterized. Bioinformatic analysis of genomic DNA sequences ofRickettsiaidentified putative lysine methyltransferases. The genes of the potential methyltransferases were synthesized, cloned, and expressed inEscherichia coli, and expressed proteins were purified by nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. The methyltransferase activities of the purified proteins were analyzed by methyl incorporation of radioactively labeledS-adenosylmethionine into recombinant fragments of OmpB. Two putative recombinant methyltransferases (rRP789 and rRP027-028) methylated recombinant OmpB fragments. The specific activity of rRP789 is 10- to 30-fold higher than that of rRP027-028. Western blot analysis using specific antibodies against trimethyl lysine showed that both rRP789 and rRP027-028 catalyzed trimethylation of recombinant OmpB fragments. Liquid chromatography-tandem mass spectrometry (LC/MS-MS) analysis showed that rRP789 catalyzed mono-, di-, and trimethylation of lysine, while rRP027-028 catalyzed exclusively trimethylation. To our knowledge, rRP789 and rRP027-028 are the first biochemically characterized lysine methyltransferases of outer membrane proteins from Gram-negative bacteria. The production and characterization of rickettsial lysine methyltransferases provide new tools to investigate the mechanism of methylation of OmpB, effects of methylation on the structure and function of OmpB, and development of methylated OmpB-based diagnostic assays and vaccine candidates.

scholarly journals Anaplasma phagocytophilum Outer Membrane Protein A Interacts with Sialylated Glycoproteins To Promote Infection of Mammalian Host Cells

2012 ◽  
Vol 80 (11) ◽  
pp. 3748-3760 ◽  
Author(s):  
Nore Ojogun ◽  
Amandeep Kahlon ◽  
Stephanie A. Ragland ◽  
Matthew J. Troese ◽  
Juliana E. Mastronunzio ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Anaplasma Phagocytophilum ◽  
Protein A ◽  
Host Cells ◽  
Mammalian Host ◽  
Content Type ◽  
Outer Membrane Protein A ◽  
Sialylated Glycoproteins

ABSTRACTAnaplasma phagocytophilumis the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis (HGA).A. phagocytophilumbinding to sialyl Lewis x (sLex) and other sialylated glycans that decorate P selectin glycoprotein 1 (PSGL-1) and other glycoproteins is critical for infection of mammalian host cells. Here, we demonstrate the importance ofA. phagocytophilumouter membrane protein A (OmpA) APH_0338 in infection of mammalian host cells. OmpA is transcriptionally induced during transmission feeding ofA. phagocytophilum-infected ticks on mice and is upregulated during invasion of HL-60 cells. OmpA is presented on the pathogen's surface. Sera from HGA patients and experimentally infected mice recognize recombinant OmpA. Pretreatment ofA. phagocytophilumorganisms with OmpA antiserum reduces their abilities to infect HL-60 cells. The OmpA N-terminal region is predicted to contain the protein's extracellular domain. GlutathioneS-transferase (GST)-tagged versions of OmpA and OmpA amino acids 19 to 74 (OmpA19-74) but not OmpA75-205bind to, and competitively inhibitA. phagocytophiluminfection of, host cells. Pretreatment of host cells with sialidase or trypsin reduces or nearly eliminates, respectively, GST-OmpA adhesion. Therefore, OmpA interacts with sialylated glycoproteins. This study identifies the firstA. phagocytophilumadhesin-receptor pair and delineates the region of OmpA that is critical for infection.

scholarly journals The Helicobacter pylori Autotransporter ImaA (HP0289) Modulates the Immune Response and Contributes to Host Colonization

2012 ◽  
Vol 80 (7) ◽  
pp. 2286-2296 ◽  
Author(s):  
William E. Sause ◽  
Andrea R. Castillo ◽  
Karen M. Ottemann
Keyword(s):  
Immune Response ◽  
Helicobacter Pylori ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Dependent Manner ◽  
Wild Type ◽  
Content Type ◽  
H Pylori ◽  
Murine Infections

ABSTRACTThe human pathogenHelicobacter pyloriemploys a diverse collection of outer membrane proteins to colonize, persist, and drive disease within the acidic gastric environment. In this study, we sought to elucidate the function of the host-induced geneHP0289, which encodes an uncharacterized outer membrane protein. We first generated an isogenicH. pylorimutant that lacksHP0289and found that the mutant has a colonization defect in single-strain infections and is greatly outcompeted in mouse coinfection experiments with wild-typeH. pylori. Furthermore, we used protease assays and biochemical fractionation coupled with an HP0289-targeted peptide antibody to verify that the HP0289 protein resides in the outer membrane. Our previous findings showed that theHP0289promoter is upregulated in the mouse stomach, and here we demonstrate thatHP0289expression is induced under acidic conditions in an ArsRS-dependent manner. Finally, we have shown that theHP0289mutant induces greater expression of the chemokine interleukin-8 (IL-8) and the cytokine tumor necrosis factor alpha (TNF-α) in gastric carcinoma cells (AGS). Similarly, transcription of the IL-8 homolog keratinocyte-derived chemokine (KC) is elevated in murine infections with the HP0289 mutant than in murine infections with wild-typeH. pylori. On the basis of this phenotype, we renamed HP0289 ImaA forimmunomodulatoryautotransporter protein. Our work has revealed that genes inducedin vivoplay an important role inH. pyloripathogenesis. Specifically, the outer membrane protein ImaA modulates a component of the host inflammatory response, and thus may allowH. pylorito fine tune the host immune response based on ImaA expression.

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