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An RND-type efflux system in Borrelia burgdorferi is involved in virulence and resistance to antimicrobial compounds.

Bunikis I, Denker K, Ostberg Y, Andersen C, Benz R, Bergström S - PLoS Pathog. (2008)

Bottom Line: The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure.We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB.We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Umeå University, Umeå, Sweden.

ABSTRACT
Borrelia burgdorferi is remarkable for its ability to thrive in widely different environments due to its ability to infect various organisms. In comparison to enteric Gram-negative bacteria, these spirochetes have only a few transmembrane proteins some of which are thought to play a role in solute and nutrient uptake and excretion of toxic substances. Here, we have identified an outer membrane protein, BesC, which is part of a putative export system comprising the components BesA, BesB and BesC. We show that BesC, a TolC homolog, forms channels in planar lipid bilayers and is involved in antibiotic resistance. A besC knockout was unable to establish infection in mice, signifying the importance of this outer membrane channel in the mammalian host. The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure. We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB. We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.

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Related in: MedlinePlus

Schematic representation of the putative besB-besA-besC operon in Borrelia burgdorferi genome and results from the reverse transcription PCR.Lane 1 and 8; molecular weight markers, lane 2; PCR reaction without template using primer pair c2 and c3 served as negative control, lane 3; primer pair c2 and c3 in ordinary PCR reaction using RNA as template served as negative control for DNA contamination, lane 4; primer pair b1 and a1, lane 5; primer pair a2 and c1, lane 6; primer pair b1 and c1, lane 7; primer pair c2 and c3 served as positive control.
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ppat-1000009-g001: Schematic representation of the putative besB-besA-besC operon in Borrelia burgdorferi genome and results from the reverse transcription PCR.Lane 1 and 8; molecular weight markers, lane 2; PCR reaction without template using primer pair c2 and c3 served as negative control, lane 3; primer pair c2 and c3 in ordinary PCR reaction using RNA as template served as negative control for DNA contamination, lane 4; primer pair b1 and a1, lane 5; primer pair a2 and c1, lane 6; primer pair b1 and c1, lane 7; primer pair c2 and c3 served as positive control.

Mentions: According to the B. burgdorferi genome sequence, the genes besB (bb0140), besA (bb0141) and besC (bb0142) are oriented in the same direction, and separated by 18 and 8 bp, respectively. In order to determine whether the three genes are transcribed as a single transcript, we analyzed RNA from low-passage strain B31 by RT-PCR using primers specific for different genes (Table S1). The primer pair spanning junction of besB and besA amplified a fragment (lane 4 in Figure 1) demonstrating that both genes are located on the same transcript. Similar results were obtained for primer pair spanning junction of besA and besC genes (lane 5 in Figure 1). Therefore we conclude that all three genes are transcribed together. This was further confirmed by amplifying a product spanning besB, besA and besC using primers specific for besB and besC (lane 6 in Figure 1). Negative control reactions, in which reverse transcriptase was omitted, yielded no such product (lane 3 in Figure 1), indicating that the products were derived from RNA rather than contaminating DNA. Additionally, besC-specific primers were used as a positive control (lane 7 in Figure 1). These data show that besB, besA and besC are transcribed as a single transcript.


An RND-type efflux system in Borrelia burgdorferi is involved in virulence and resistance to antimicrobial compounds.

Bunikis I, Denker K, Ostberg Y, Andersen C, Benz R, Bergström S - PLoS Pathog. (2008)

Schematic representation of the putative besB-besA-besC operon in Borrelia burgdorferi genome and results from the reverse transcription PCR.Lane 1 and 8; molecular weight markers, lane 2; PCR reaction without template using primer pair c2 and c3 served as negative control, lane 3; primer pair c2 and c3 in ordinary PCR reaction using RNA as template served as negative control for DNA contamination, lane 4; primer pair b1 and a1, lane 5; primer pair a2 and c1, lane 6; primer pair b1 and c1, lane 7; primer pair c2 and c3 served as positive control.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2279261&req=5

ppat-1000009-g001: Schematic representation of the putative besB-besA-besC operon in Borrelia burgdorferi genome and results from the reverse transcription PCR.Lane 1 and 8; molecular weight markers, lane 2; PCR reaction without template using primer pair c2 and c3 served as negative control, lane 3; primer pair c2 and c3 in ordinary PCR reaction using RNA as template served as negative control for DNA contamination, lane 4; primer pair b1 and a1, lane 5; primer pair a2 and c1, lane 6; primer pair b1 and c1, lane 7; primer pair c2 and c3 served as positive control.
Mentions: According to the B. burgdorferi genome sequence, the genes besB (bb0140), besA (bb0141) and besC (bb0142) are oriented in the same direction, and separated by 18 and 8 bp, respectively. In order to determine whether the three genes are transcribed as a single transcript, we analyzed RNA from low-passage strain B31 by RT-PCR using primers specific for different genes (Table S1). The primer pair spanning junction of besB and besA amplified a fragment (lane 4 in Figure 1) demonstrating that both genes are located on the same transcript. Similar results were obtained for primer pair spanning junction of besA and besC genes (lane 5 in Figure 1). Therefore we conclude that all three genes are transcribed together. This was further confirmed by amplifying a product spanning besB, besA and besC using primers specific for besB and besC (lane 6 in Figure 1). Negative control reactions, in which reverse transcriptase was omitted, yielded no such product (lane 3 in Figure 1), indicating that the products were derived from RNA rather than contaminating DNA. Additionally, besC-specific primers were used as a positive control (lane 7 in Figure 1). These data show that besB, besA and besC are transcribed as a single transcript.

Bottom Line: The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure.We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB.We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Umeå University, Umeå, Sweden.

ABSTRACT
Borrelia burgdorferi is remarkable for its ability to thrive in widely different environments due to its ability to infect various organisms. In comparison to enteric Gram-negative bacteria, these spirochetes have only a few transmembrane proteins some of which are thought to play a role in solute and nutrient uptake and excretion of toxic substances. Here, we have identified an outer membrane protein, BesC, which is part of a putative export system comprising the components BesA, BesB and BesC. We show that BesC, a TolC homolog, forms channels in planar lipid bilayers and is involved in antibiotic resistance. A besC knockout was unable to establish infection in mice, signifying the importance of this outer membrane channel in the mammalian host. The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure. We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB. We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.

Show MeSH
Related in: MedlinePlus