Major outer membrane protein of Legionella pneumophila carries a species-specific epitope.

Legionella pneumophila is a facultative intracellular bacterial pathogen that parasitizes human monocytes and alveolar macrophages. Previous studies from this laboratory have shown that monocyte complement receptors CR1 and CR3 and complement component C3 in serum mediate L. pneumophila phagocytosis. In this study, we have explored C3 fixation to L. pneumophila. We developed a whole-cell enzyme-linked immunosorbent assay (ELISA) to measure C3 fixation to the bacterial surface. By this assay, C3 fixes to L. pneumophila that are opsonized in fresh nonimmune serum, and C3 fixation takes place via the alternative pathway of complement activation. Immunoblot analysis of opsonized L. pneumophila indicated that C3 fixes selectively to specific acceptor molecules of L. pneumophila. Consistent with this, when nitrocellulose blots of whole L. pneumophila or bacterial components are incubated in fresh nonimmune serum, C3 fixes exclusively to the major outer membrane protein (MOMP) of L. pneumophila, a porin; C3 does not fix to L. pneumophila LPS on these blots. To further explore the role of MOMP in C3 fixation and phagocytosis, we reconstituted purified MOMP into liposomes. By the ELISA, MOMP-liposomes, but not plain liposomes lacking MOMP, avidly fix C3. Consistent with a dominant role for MOMP in C3 fixation, MOMP-liposomes form a C3 complex of the same apparent molecular weight as whole L. pneumophila in nonimmune serum. Opsonized radioiodinated MOMP-liposomes avidly adhere to monocytes, and adherence is dose dependent upon serum. By electron microscopy, opsonized MOMP-liposomes are efficiently phagocytized by human monocytes, and phagocytosis takes place by a conventional appearing form of phagocytosis. This study demonstrates that C3 fixes selectively to the MOMP of L. pneumophila, and that, in the presence of nonimmune serum, MOMP can mediate phagocytosis of liposomes and, potentially, phagocytosis of intact L. pneumophila by human monocytes.

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Legionella pneumophila DNA coding for major outer membrane protein; serogroup-1 Philadelphia strain 29 kDa recombinant major outer membrane protein production; DNA sequence; application in diagnosis and recombinant vaccine

Vaccine ◽

10.1016/s0264-410x(96)90018-3 ◽

1996 ◽

Vol 14 (5) ◽

pp. 458

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Dna Sequence ◽

Outer Membrane Protein ◽

Legionella Pneumophila ◽

Protein Production ◽

Recombinant Vaccine ◽

Major Outer Membrane Protein ◽

Dna Coding

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Chlamydia pneumoniae Major Outer Membrane Protein Is a Surface-Exposed Antigen That Elicits Antibodies Primarily Directed against Conformation-Dependent Determinants

Infection and Immunity ◽

10.1128/iai.69.5.3082-3091.2001 ◽

2001 ◽

Vol 69 (5) ◽

pp. 3082-3091 ◽

Cited By ~ 45

Author(s):

Katerina Wolf ◽

Elizabeth Fischer ◽

David Mead ◽

Guangming Zhong ◽

Roseanna Peeling ◽

...

Keyword(s):

Monoclonal Antibody ◽

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Chlamydia Pneumoniae ◽

Major Outer Membrane Protein ◽

Mass Spectrometry Analysis ◽

Antigenic Determinants ◽

Specific Monoclonal Antibody ◽

Species Specific

ABSTRACT The major outer membrane protein (MOMP) of Chlamydia trachomatis serovariants is known to be an immunodominant surface antigen. Moreover, it is known that the C. trachomatis MOMP elicits antibodies that recognize both linear and conformational antigenic determinants. In contrast, it has been reported that the MOMP of Chlamydia pneumoniae is not surface exposed and is immunorecessive. We hypothesized that the discrepancies betweenC. trachomatis and C. pneumoniae MOMP exposure on intact chlamydiae and immunogenic properties might be because the focus of the host's immune response is directed to conformational epitopes of the C. pneumoniae MOMP. We therefore conducted studies aimed at defining the surface exposure of MOMP and the conformational dominance of MOMP antibodies. We present here a description of C. pneumoniaespecies-specific monoclonal antibody (MAb), GZD1E8, which recognizes a conformational epitope on the surface of C. pneumoniae. This MAb is potent in the neutralization ofC. pneumoniae infectivity in vitro. Another previously described C. pneumoniaespecies-specific monoclonal antibody, RR-402, displayed very similar characteristics. However, the antigenic determinant recognized by RR-402 has yet to be identified. We show by immunoprecipitation ofC. pneumoniae with GZD1E8 and RR-402 MAbs and by mass spectrometry analysis of immunoprecipitated proteins that both antibodies GZD1E8 and RR-402 recognize the MOMP of C. pneumoniae and that this protein is localized on the surface of the organism. We also show that human sera fromC. pneumoniae-positive donors consistently recognize the MOMP by immunoprecipitation, indicating that the MOMP ofC. pneumoniae is an immunogenic protein. These findings have potential implications for both C. pneumoniae vaccine and diagnostic assay development.

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Major Outer Membrane Protein from Legionella pneumophila Inhibits Phagocytosis but Enhances Chemotaxis of RAW 264.7 Macrophages by Regulating the FOXO1/Coronin-1 Axis

Journal of Immunology Research ◽

10.1155/2021/9409777 ◽

2021 ◽

Vol 2021 ◽

pp. 1-11

Author(s):

Zehui Yang ◽

Yingying Chen ◽

Qiang Zhang ◽

Xiaodong Chen ◽

Ze Deng

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Legionella Pneumophila ◽

Major Outer Membrane Protein ◽

Raw 264.7 ◽

Dependent Manner ◽

Raw 264.7 Macrophages ◽

Expression Levels ◽

Raw 264.7 Macrophage

Legionella pneumophila is an intracellular pathogen that can cause Legionnaire’s disease by invading alveolar epithelial cells and macrophages. The major outer membrane protein (MOMP) plays an important role in the interaction between bacteria and host cells. However, the role of MOMP in the process of L. pneumophila invasion of macrophages and its working mechanism remain unknown. We aimed to explore the effects of MOMP on phagocytosis and chemotaxis of RAW 264.7 macrophages. The chemotactic activity, toxicity, and phagocytosis of RAW 264.7 cocultured with different concentrations of MOMP were determined by Transwell, CCK-8, and neutral red uptake assays, respectively. Target genes were detected by double-luciferase and pull down assays. qRT-PCR and Western blot were performed to analyze the expression of several important proteins involved in the immune response pathway, including coronin-1, interleukins (IL-10), forkhead transcription factor 1 (FOXO1), nucleotide-binding oligomerization domain protein (NOD) 1, NOD2, and receptor-interacting protein (RIP) 2. After coculturing with MOMP, cytological observation indicated a decrease of phagocytosis and a marked increase of chemotaxis in RAW 264.7 macrophages. The phagocytosis degree of RAW 264.7 macrophage varied with the concentration gradient of MOMP in a time-dependent manner. MOMP could increase the expression levels of MCP-1, IL-10, NOD2, and RIP2 and decrease the expression levels of FOXO1 and coronin-1 in cell culture supernatants. In addition, we found that FOXO1 could promote its transcription by binding to the promoter of coronin-1. The results of the present study suggested that MOMP could inhibit phagocytosis and facilitate chemotaxis of RAW 264.7 macrophage, which might be associated with the FOXO1/coronin-1 axis.

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