Dataset Information

Identification of the essential Brucella melitensis porin Omp2b as a suppressor of Bax-induced cell death in yeast in a genome-wide screening.

ABSTRACT:

Background

Inhibition of apoptosis is one of the mechanisms selected by numerous intracellular pathogenic bacteria to control their host cell. Brucellae, which are the causative agent of a worldwide zoonosis, prevent apoptosis of infected cells, probably to support survival of their replication niche.

Methodology/principal findings

In order to identify Brucella melitensis anti-apoptotic effector candidates, we performed a genome-wide functional screening in yeast. The B. melitensis ORFeome was screened to identify inhibitors of Bax-induced cell death in S. cerevisiae. B. melitensis porin Omp2b, here shown to be essential, prevents Bax lethal effect in yeast, unlike its close paralog Omp2a. Our results based on Omp2b size variants characterization suggest that signal peptide processing is required for Omp2b effect in yeast.

Conclusion/significance

We report here the first application to a bacterial genome-wide library of coding sequences of this "yeast-rescue" screening strategy, previously used to highlight several new apoptosis regulators. Our work provides B. melitensis proteins that are candidates for an anti-apoptotic function, and can be tested in mammalian cells in the future. Hypotheses on possible molecular mechanisms of Bax inhibition by the B. melitensis porin Omp2b are discussed.

SUBMITTER: Laloux G

PROVIDER: S-EPMC2952587 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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Publications

Identification of the essential Brucella melitensis porin Omp2b as a suppressor of Bax-induced cell death in yeast in a genome-wide screening.

Laloux Géraldine G Deghelt Michaël M de Barsy Marie M Letesson Jean-Jacques JJ De Bolle Xavier X

PloS one 20101011 10

<h4>Background</h4>Inhibition of apoptosis is one of the mechanisms selected by numerous intracellular pathogenic bacteria to control their host cell. Brucellae, which are the causative agent of a worldwide zoonosis, prevent apoptosis of infected cells, probably to support survival of their replication niche.<h4>Methodology/principal findings</h4>In order to identify Brucella melitensis anti-apoptotic effector candidates, we performed a genome-wide functional screening in yeast. The B. melitensi ...[more]

PMID: 20949000

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Project description:The ongoing antibiotic-resistance crisis is becoming a global problem affecting public health. Urgent efforts are required to design novel therapeutics against pathogenic bacterial species. Brucella melitensis is an etiological agent of brucellosis, which mostly affects sheep and goats but several cases have also been reported in cattle, water buffalo, yaks and dogs. Infected animals also represent the major source of infection for humans. Development of safer and effective vaccines for brucellosis remains a priority to support disease control and eradication in animals and to prevent infection to humans. In this research study, we designed an in-silico multi-epitopes vaccine for B. melitensis using computational approaches. The pathogen core proteome was screened for good vaccine candidates using subtractive proteomics, reverse vaccinology and immunoinformatic tools. In total, 10 proteins: catalase; siderophore ABC transporter substrate-binding protein; pyridoxamine 5'-phosphate oxidase; superoxide dismutase; peptidylprolyl isomerase; superoxide dismutase family protein; septation protein A; hypothetical protein; binding-protein-dependent transport systems inner membrane component; and 4-hydroxy-2-oxoheptanedioate aldolase were selected for epitopes prediction. To induce cellular and antibody base immune responses, the vaccine must comprise both B and T-cells epitopes. The epitopes were next screened for antigenicity, allergic nature and water solubility and the probable antigenic, non-allergic, water-soluble and non-toxic nine epitopes were shortlisted for multi-epitopes vaccine construction. The designed vaccine construct comprises 274 amino acid long sequences having a molecular weight of 28.14 kDa and instability index of 27.62. The vaccine construct was further assessed for binding efficacy with immune cell receptors. Docking results revealed that the designed vaccine had good binding potency with selected immune cell receptors. Furthermore, vaccine-MHC-I, vaccine-MHC-II and vaccine-TLR-4 complexes were opted based on a least-binding energy score of -5.48 kcal/mol, 0.64 kcal/mol and -2.69 kcal/mol. Those selected were then energy refined and subjected to simulation studies to understand dynamic movements of the docked complexes. The docking results were further validated through MMPBSA and MMGBSA analyses. The MMPBSA calculated -235.18 kcal/mol, -206.79 kcal/mol, and -215.73 kcal/mol net binding free energy, while MMGBSA estimated -259.48 kcal/mol, -206.79 kcal/mol and -215.73 kcal/mol for TLR-4, MHC-I and MHC-II complexes, respectively. These findings were validated by water-swap and entropy calculations. Overall, the designed vaccine construct can evoke proper immune responses and the construct could be helpful for experimental researchers in formulation of a protective vaccine against the targeted pathogen for both animal and human use.

Dataset Information

Identification of the essential Brucella melitensis porin Omp2b as a suppressor of Bax-induced cell death in yeast in a genome-wide screening.

Background

Methodology/principal findings

Conclusion/significance

Publications

Identification of the essential Brucella melitensis porin Omp2b as a suppressor of Bax-induced cell death in yeast in a genome-wide screening.

Similar Datasets

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