scholarly journals Recombinant Protein Truncation Strategy for Inducing Bactericidal Antibodies to the Macrophage Infectivity Potentiator Protein of Neisseria meningitidis and Circumventing Potential Cross-Reactivity with Human FK506-Binding Proteins

2014 ◽  
Vol 83 (2) ◽  
pp. 730-742 ◽  
Author(s):  
Magdalena K. Bielecka ◽  
Nathalie Devos ◽  
Mélanie Gilbert ◽  
Miao-Chiu Hung ◽  
Vincent Weynants ◽  
...  
Keyword(s):  
Neisseria Meningitidis ◽  
Bactericidal Activity ◽  
Binding Proteins ◽  
Leader Peptide ◽  
Type I ◽  
Globular Domain ◽  
His Tag ◽  
Macrophage Infectivity Potentiator ◽  
Fk506 Binding Proteins ◽  
Macrophage Infectivity

A recombinant macrophage infectivity potentiator (rMIP) protein ofNeisseria meningitidisinduces significant serum bactericidal antibody production in mice and is a candidate meningococcal vaccine antigen. However, bioinformatics analysis of MIP showed some amino acid sequence similarity to human FK506-binding proteins (FKBPs) in residues 166 to 252 located in the globular domain of the protein. To circumvent the potential concern over generating antibodies that could recognize human proteins, we immunized mice with recombinant truncated type I rMIP proteins that lacked the globular domain and the signal leader peptide (LP) signal sequence (amino acids 1 to 22) and contained the His purification tag at either the N or C terminus (C-term). The immunogenicity of truncated rMIP proteins was compared to that of full (i.e., full-length) rMIP proteins (containing the globular domain) with either an N- or C-terminal His tag and with or without the LP sequence. By comparing the functional murine antibody responses to these various constructs, we determined that C-term His truncated rMIP (−LP) delivered in liposomes induced high levels of antibodies that bound to the surface of wild-type but not Δmipmutant meningococci and showed bactericidal activity against homologous type I MIP (median titers of 128 to 256) and heterologous type II and III (median titers of 256 to 512) strains, thereby providing at least 82% serogroup B strain coverage. In contrast, in constructs lacking the LP, placement of the His tag at the N terminus appeared to abrogate bactericidal activity. The strategy used in this study would obviate any potential concerns regarding the use of MIP antigens for inclusion in bacterial vaccines.

scholarly journals The Neisseria meningitidis Macrophage Infectivity Potentiator Protein Induces Cross-Strain Serum Bactericidal Activity and Is a Potential Serogroup B Vaccine Candidate

2011 ◽  
Vol 79 (9) ◽  
pp. 3784-3791 ◽  
Author(s):  
Miao-Chiu Hung ◽  
Omar Salim ◽  
Jeannette N. Williams ◽  
John E. Heckels ◽  
Myron Christodoulides
Keyword(s):  
Neisseria Meningitidis ◽  
Bactericidal Activity ◽  
Legionella Pneumophila ◽  
Type I ◽  
Content Type ◽  
Serum Bactericidal Activity ◽  
Monophosphoryl Lipid A ◽  
Macrophage Infectivity Potentiator ◽  
Sequence Types ◽  
Macrophage Infectivity

ABSTRACTA gene encoding a 29-kDa protein fromNeisseria meningitidisserogroup B strain MC58 with homology to the macrophage infectivity potentiator (MIP) protein ofLegionella pneumophilawas cloned and expressed inEscherichia coli, and the purified soluble recombinant protein (rMIP) was used for immunization studies. Analysis of the predicted amino acid sequences of MIP from 13 well-characterized meningococcal strains, isolated from carriers or patients and differing in serogroup, serotype, and subtype, showed that the protein was highly conserved (98 to 100%), with only three distinct sequence types (designated I, II, and III) found. Western blotting showed that the MIP protein was expressed at similar levels by all of these strains. Immunization of mice with type I MC58 rMIP in detergent micelles and liposomes containing monophosphoryl lipid A (MPLA) induced high levels of surface-reactive antibodies with serum bactericidal activity (SBA) titers of 1/1,024 against the homologous strain. Bactericidal antibodies were also induced with the protein in saline alone and liposomes alone (titers, 1/128) but not following adsorption to Al(OH)3. Significantly, antisera raised against type I rMIP administered in saline or liposomes killed strains of heterologous sequence types II and III with similar SBA titers (1/128 to 1/256). Taken together, these findings suggest that rMIP can provide cross-strain protection against meningococci and should be considered a potential antigen for inclusion in new vaccines against meningococcal infection.

Liposomes or traditional adjuvants: induction of bactericidal activity by the macrophage infectivity potentiator protein (Mip) ofNeisseria meningitidis

Apmis ◽  
2017 ◽  
Vol 125 (8) ◽  
pp. 725-731 ◽  
Author(s):  
Liliana Costoya ◽  
Juan Marzoa ◽  
Carlos Ferreirós ◽  
Maria Teresa Criado
Keyword(s):  
Bactericidal Activity ◽  
Macrophage Infectivity Potentiator ◽  
Macrophage Infectivity

scholarly journals The Adhesin Complex Protein (ACP) of Neisseria meningitidis Is a New Adhesin with Vaccine Potential

mBio ◽  
2013 ◽  
Vol 4 (2) ◽  
Author(s):  
Miao-Chiu Hung ◽  
John E. Heckels ◽  
Myron Christodoulides
Keyword(s):  
Neisseria Meningitidis ◽  
Bactericidal Activity ◽  
Human Cells ◽  
Amino Acid Sequences ◽  
Target Antigen ◽  
Type I ◽  
Surface Binding ◽  
Content Type ◽  
Complex Protein ◽  
Sequence Types

ABSTRACTTheacpgene encoding the 13-kDa adhesin complex protein (ACP) fromNeisseria meningitidisserogroup B strain MC58 was cloned and expressed inEscherichia coli, and the purified recombinant ACP (rACP) was used for immunization studies. Analysis of the ACP amino acid sequences from 13 meningococcal strains, isolated from patients and colonized individuals, and 178 strains in the Bacterial Isolate Genome Sequence (BIGS) database showed the presence of only three distinct sequence types (I, II, and III) with high similarity (>98%). Immunization of mice with type I rACP in detergent micelles and liposomes and in saline solution alone induced high levels of serum bactericidal activity (SBA; titers of 1/512) against the homologous strain MC58 and killed strains of heterologous sequence types II and III with similar SBA titers (1/128 to 1/512). Levels of expression of type I, II, or III ACP by different meningococcal strains were similar. ACP functioned as an adhesin, as demonstrated by reduced adherence ofacpknockout (MC58 ΔACP) meningococci to human cellsin vitroand the direct surface binding of rACP and by the ability of anti-rACP sera to inhibit adherence of wild-type bacteria. ACP also mediated the invasion of noncapsular meningococci into human epithelial cells, but it was not a particularly impressive invasin, as the internalized bacterial numbers were low. In summary, the newly identified ACP protein is an adhesin that induces cross-strain bactericidal activity and is therefore an attractive target antigen for incorporation into the next generation of serogroup B meningococcal vaccines.IMPORTANCEInfections caused byNeisseria meningitidisserogroup B are still significant causes of mortality and morbidity worldwide, and broadly protective vaccines of defined antigen composition are not yet licensed. Here, we describe the properties of the adhesin complex protein (ACP), which we demonstrate is a newly recognized molecule that is highly conserved and expressed to similar levels in meningococci and facilitates meningococcal interactions with human cells. We also report that a recombinant ACP protein vaccine induces murine antibodies that significantly kill meningococci expressing different ACP. Taken together, these properties demonstrate that ACP merits serious consideration as a component of a broadly protective vaccine against serogroup B meningococci.

scholarly journals Color-tuning of natural variants of heliorhodopsin

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Se-Hwan Kim ◽  
Kimleng Chuon ◽  
Shin-Gyu Cho ◽  
Ahreum Choi ◽  
Seanghun Meas ◽  
...  
Keyword(s):  
3D Structure ◽  
Site Directed Mutagenesis ◽  
Type I ◽  
Transmembrane Helices ◽  
Color Tuning ◽  
Tuning Mechanism ◽  
Spectral Changes ◽  
His Tag ◽  
Natural Variants ◽  
Retinal Chromophore

AbstractMicrobial rhodopsins are distributed through many microorganisms. Heliorhodopsins are newly discovered but have an unclear function. They have seven transmembrane helices similar to type-I and type-II rhodopsins, but they are different in that the N-terminal region of heliorhodopsin is cytoplasmic. We chose 13 representative heliorhodopsins from various microorganisms, expressed and purified with an N-terminal His tag, and measured the absorption spectra. The 13 natural variants had an absorption maximum (λmax) in the range 530–556 nm similar to proteorhodopsin (λmax = 490–525 nm). We selected several candidate residues that influence rhodopsin color-tuning based on sequence alignment and constructed mutants via site-directed mutagenesis to confirm the spectral changes. We found two important residues located near retinal chromophore that influence λmax. We also predict the 3D structure via homology-modeling of Thermoplasmatales heliorhodopsin. The results indicate that the color-tuning mechanism of type-I rhodopsin can be applied to understand the color-tuning of heliorhodopsin.

scholarly journals Inhibition of FK506 Binding Proteins Reduces  -Synuclein Aggregation and Parkinson's Disease-Like Pathology

2010 ◽  
Vol 30 (7) ◽  
pp. 2454-2463 ◽  
Author(s):  
M. Gerard ◽  
A. Deleersnijder ◽  
V. Daniels ◽  
S. Schreurs ◽  
S. Munck ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Binding Proteins ◽  
Fk506 Binding Proteins

scholarly journals MisR/MisS Two-Component Regulon in Neisseria meningitidis

2007 ◽  
Vol 76 (2) ◽  
pp. 704-716 ◽  
Author(s):  
Yih-Ling Tzeng ◽  
Charlene M. Kahler ◽  
Xinjian Zhang ◽  
David S. Stephens
Keyword(s):  
Neisseria Meningitidis ◽  
Target Genes ◽  
Inner Core ◽  
Consensus Sequence ◽  
Meningococcal Infection ◽  
Type I ◽  
Mobility Shift ◽  
Iron Assimilation ◽  
Wide Range ◽  
Two Component

ABSTRACT Two-component regulatory systems are involved in processes important for bacterial pathogenesis. Inactivation of the misR/misS system in Neisseria meningitidis results in the loss of phosphorylation of the lipooligosaccharide inner core and causes attenuation in a mouse model of meningococcal infection. One hundred seventeen (78 up-regulated and 39 down-regulated) potential regulatory targets of the MisR/MisS (MisR/S) system were identified by transcriptional profiling of the NMBmisR mutant and the parental wild-type meningococcal strain NMB. The regulatory effect was further confirmed in a subset of target genes by quantitative real-time PCR and β-galactosidase transcriptional fusion reporter assays. The MisR regulon includes genes encoding proteins necessary for protein folding in the bacterial cytoplasm and periplasm, transcriptional regulation, metabolism, iron assimilation, and type I protein transport. Mutation in the MisR/S system caused increased sensitivity to oxidative stress and also resulted in decreased susceptibility to complement-mediated killing by normal human serum. To identify the direct targets of MisR regulation, electrophoretic mobility shift assays were carried out using purified MisR-His6 protein. Among 22 genes examined, misR directly interacted with 14 promoter regions. Six promoters were further investigated by DNase I protection assays, and a MisR-binding consensus sequence was proposed. Thus, the direct regulatory targets of MisR and the minimal regulon of the meningococcal MisR/S two-component signal transduction system were characterized. These data indicate that the MisR/S system influences a wide range of biological functions in N. meningitidis either directly or via intermediate regulators.

Inhibitors of macrophage infectivity potentiator-like PPIases affect neisserial and chlamydial pathogenicity

2016 ◽  
Vol 48 (4) ◽  
pp. 401-408 ◽  
Author(s):  
Anastasija Reimer ◽  
Florian Seufert ◽  
Matthias Weiwad ◽  
Jutta Ebert ◽  
Nicole M. Bzdyl ◽  
...  
Keyword(s):  
Macrophage Infectivity Potentiator ◽  
Macrophage Infectivity
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