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YqiC of Salmonella enterica serovar Typhimurium is a membrane fusogenic protein required for mice colonization.

Carrica MC, Craig PO, García-Angulo VA, Aguirre A, García-Véscovi E, Goldbaum FA, Cravero SL - BMC Microbiol. (2011)

Bottom Line: We found that YqiC shares biophysical and biochemical properties with Brucella abortus BMFP, the only previously characterized member of this group, such as a high alpha helix content, a coiled-coil domain involved in trimerization and a membrane fusogenic activity in vitro.In addition, we demonstrated that YqiC localizes at cytoplasmic and membrane subcellular fractions, that a S.This work firstly demonstrates the importance of a COG2960 member for pathogen-host interaction, and suggests a common function conserved among members of this group.

View Article: PubMed Central - HTML - PubMed

Affiliation: Instituto de Biotecnología, CICVyA-INTA Castelar, Los Reseros y Las Cabañas s/n, Buenos Aires, Argentina. mcarrica@leloir.org.ar

ABSTRACT

Background: Salmonella enterica serovar Typhimurium is an intracellular bacterial pathogen which can colonize a variety of hosts, including human, causing syndromes that vary from gastroenteritis and diarrhea to systemic disease.

Results: In this work we present structural information as well as insights into the in vivo function of YqiC, a 99-residue protein of S. Typhimurium, which belongs to the cluster of the orthologous group 2960 (COG2960). We found that YqiC shares biophysical and biochemical properties with Brucella abortus BMFP, the only previously characterized member of this group, such as a high alpha helix content, a coiled-coil domain involved in trimerization and a membrane fusogenic activity in vitro. In addition, we demonstrated that YqiC localizes at cytoplasmic and membrane subcellular fractions, that a S. Typhimurium yqiC deficient strain had a severe attenuation in virulence in the murine model when inoculated both orally and intraperitoneally, and was impaired to replicate at physiological and high temperatures in vitro, although it was still able to invade and replicate inside epithelial and macrophages cell lines.

Conclusion: This work firstly demonstrates the importance of a COG2960 member for pathogen-host interaction, and suggests a common function conserved among members of this group.

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Related in: MedlinePlus

Subcellular localization of YqiC. Whole-cell lysate of S. Typhimurium was fractionated by ultracentrifugation. Samples of the cell lysate (L), the supernatant (S) and the sedimented membrane fraction (M) were analyzed by immunoblotting with anti-YqiC and anti-MBP antiserum. Antibodies against the soluble MBP protein [10] was used as a control for the membrane fraction contamination.
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Figure 4: Subcellular localization of YqiC. Whole-cell lysate of S. Typhimurium was fractionated by ultracentrifugation. Samples of the cell lysate (L), the supernatant (S) and the sedimented membrane fraction (M) were analyzed by immunoblotting with anti-YqiC and anti-MBP antiserum. Antibodies against the soluble MBP protein [10] was used as a control for the membrane fraction contamination.

Mentions: To determine the subcellular localization of YqiC, we performed a mechanical lysis fractionation procedure. A wild type S. Typhimurium culture grown to late log phase was harvested by centrifugation, mechanically disrupted and fractionated by ultracentrifugation. This procedure allows for the separation of bacterial proteins into two fractions: the supernatant, which contains cytoplasmic and periplasmic proteins, and the pellet fraction, which contains the inner and outer membrane proteins. Fractions were then analyzed by immunoblotting using an anti-YqiC polyclonal antibody. YqiC was localized in the two fractions, although lower levels of YqiC were found in the membrane fraction (Figure 4). This result indicated that YqiC is both soluble and membrane associated inside the cell. As a control, we used an antibody against the periplasmic protein MBP [10], which was only detected in the supernatant fraction.


YqiC of Salmonella enterica serovar Typhimurium is a membrane fusogenic protein required for mice colonization.

Carrica MC, Craig PO, García-Angulo VA, Aguirre A, García-Véscovi E, Goldbaum FA, Cravero SL - BMC Microbiol. (2011)

Subcellular localization of YqiC. Whole-cell lysate of S. Typhimurium was fractionated by ultracentrifugation. Samples of the cell lysate (L), the supernatant (S) and the sedimented membrane fraction (M) were analyzed by immunoblotting with anti-YqiC and anti-MBP antiserum. Antibodies against the soluble MBP protein [10] was used as a control for the membrane fraction contamination.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3107778&req=5

Figure 4: Subcellular localization of YqiC. Whole-cell lysate of S. Typhimurium was fractionated by ultracentrifugation. Samples of the cell lysate (L), the supernatant (S) and the sedimented membrane fraction (M) were analyzed by immunoblotting with anti-YqiC and anti-MBP antiserum. Antibodies against the soluble MBP protein [10] was used as a control for the membrane fraction contamination.
Mentions: To determine the subcellular localization of YqiC, we performed a mechanical lysis fractionation procedure. A wild type S. Typhimurium culture grown to late log phase was harvested by centrifugation, mechanically disrupted and fractionated by ultracentrifugation. This procedure allows for the separation of bacterial proteins into two fractions: the supernatant, which contains cytoplasmic and periplasmic proteins, and the pellet fraction, which contains the inner and outer membrane proteins. Fractions were then analyzed by immunoblotting using an anti-YqiC polyclonal antibody. YqiC was localized in the two fractions, although lower levels of YqiC were found in the membrane fraction (Figure 4). This result indicated that YqiC is both soluble and membrane associated inside the cell. As a control, we used an antibody against the periplasmic protein MBP [10], which was only detected in the supernatant fraction.

Bottom Line: We found that YqiC shares biophysical and biochemical properties with Brucella abortus BMFP, the only previously characterized member of this group, such as a high alpha helix content, a coiled-coil domain involved in trimerization and a membrane fusogenic activity in vitro.In addition, we demonstrated that YqiC localizes at cytoplasmic and membrane subcellular fractions, that a S.This work firstly demonstrates the importance of a COG2960 member for pathogen-host interaction, and suggests a common function conserved among members of this group.

View Article: PubMed Central - HTML - PubMed

Affiliation: Instituto de Biotecnología, CICVyA-INTA Castelar, Los Reseros y Las Cabañas s/n, Buenos Aires, Argentina. mcarrica@leloir.org.ar

ABSTRACT

Background: Salmonella enterica serovar Typhimurium is an intracellular bacterial pathogen which can colonize a variety of hosts, including human, causing syndromes that vary from gastroenteritis and diarrhea to systemic disease.

Results: In this work we present structural information as well as insights into the in vivo function of YqiC, a 99-residue protein of S. Typhimurium, which belongs to the cluster of the orthologous group 2960 (COG2960). We found that YqiC shares biophysical and biochemical properties with Brucella abortus BMFP, the only previously characterized member of this group, such as a high alpha helix content, a coiled-coil domain involved in trimerization and a membrane fusogenic activity in vitro. In addition, we demonstrated that YqiC localizes at cytoplasmic and membrane subcellular fractions, that a S. Typhimurium yqiC deficient strain had a severe attenuation in virulence in the murine model when inoculated both orally and intraperitoneally, and was impaired to replicate at physiological and high temperatures in vitro, although it was still able to invade and replicate inside epithelial and macrophages cell lines.

Conclusion: This work firstly demonstrates the importance of a COG2960 member for pathogen-host interaction, and suggests a common function conserved among members of this group.

Show MeSH
Related in: MedlinePlus