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Inhibition of bacterial conjugation by phage M13 and its protein g3p: quantitative analysis and model.

Lin A, Jimenez J, Derr J, Vera P, Manapat ML, Esvelt KM, Villanueva L, Liu DR, Chen IA - PLoS ONE (2011)

Bottom Line: Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances.We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration.Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage), these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

View Article: PubMed Central - PubMed

Affiliation: FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts, United States of America.

ABSTRACT
Conjugation is the main mode of horizontal gene transfer that spreads antibiotic resistance among bacteria. Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances. Filamentous bacteriophages were first observed to inhibit conjugation several decades ago. Here we investigate the mechanism of inhibition and find that the primary effect on conjugation is occlusion of the conjugative pilus by phage particles. This interaction is mediated primarily by phage coat protein g3p, and exogenous addition of the soluble fragment of g3p inhibited conjugation at low nanomolar concentrations. Our data are quantitatively consistent with a simple model in which association between the pili and phage particles or g3p prevents transmission of an F plasmid encoding tetracycline resistance. We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration. Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage), these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

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Inhibition of conjugation by minor coat protein g3p from M13: Inhibition curve for conjugation in the presence of g3p-N protein (red) or BSA (blue).Line represents model fit to a binding equilibrium (Kd = 3 nM; RMSD = 0.1). Error bars indicate standard deviation from experimental replicates.
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pone-0019991-g005: Inhibition of conjugation by minor coat protein g3p from M13: Inhibition curve for conjugation in the presence of g3p-N protein (red) or BSA (blue).Line represents model fit to a binding equilibrium (Kd = 3 nM; RMSD = 0.1). Error bars indicate standard deviation from experimental replicates.

Mentions: To confirm that g3p was the causative factor, we obtained a purified soluble fragment of g3p comprising the N-terminal domains (g3p-N) [22]. Addition of this protein to the media inhibited conjugation (Figure 4B), and the inhibition could be described well as a simple binding equilibrium between protein and F+ cells, similar to that described above for the non-replicating phage particles (Figure 5). The dissociation constant of this equilibrium was found to be 3 nM. The presence of g3p-N did not alter the growth rate of cells (Table 1). Addition of bovine serum albumin to high concentration did not appreciably inhibit conjugation (Figure 4A, Figure 5).


Inhibition of bacterial conjugation by phage M13 and its protein g3p: quantitative analysis and model.

Lin A, Jimenez J, Derr J, Vera P, Manapat ML, Esvelt KM, Villanueva L, Liu DR, Chen IA - PLoS ONE (2011)

Inhibition of conjugation by minor coat protein g3p from M13: Inhibition curve for conjugation in the presence of g3p-N protein (red) or BSA (blue).Line represents model fit to a binding equilibrium (Kd = 3 nM; RMSD = 0.1). Error bars indicate standard deviation from experimental replicates.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3102678&req=5

pone-0019991-g005: Inhibition of conjugation by minor coat protein g3p from M13: Inhibition curve for conjugation in the presence of g3p-N protein (red) or BSA (blue).Line represents model fit to a binding equilibrium (Kd = 3 nM; RMSD = 0.1). Error bars indicate standard deviation from experimental replicates.
Mentions: To confirm that g3p was the causative factor, we obtained a purified soluble fragment of g3p comprising the N-terminal domains (g3p-N) [22]. Addition of this protein to the media inhibited conjugation (Figure 4B), and the inhibition could be described well as a simple binding equilibrium between protein and F+ cells, similar to that described above for the non-replicating phage particles (Figure 5). The dissociation constant of this equilibrium was found to be 3 nM. The presence of g3p-N did not alter the growth rate of cells (Table 1). Addition of bovine serum albumin to high concentration did not appreciably inhibit conjugation (Figure 4A, Figure 5).

Bottom Line: Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances.We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration.Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage), these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

View Article: PubMed Central - PubMed

Affiliation: FAS Center for Systems Biology, Harvard University, Cambridge, Massachusetts, United States of America.

ABSTRACT
Conjugation is the main mode of horizontal gene transfer that spreads antibiotic resistance among bacteria. Strategies for inhibiting conjugation may be useful for preserving the effectiveness of antibiotics and preventing the emergence of bacterial strains with multiple resistances. Filamentous bacteriophages were first observed to inhibit conjugation several decades ago. Here we investigate the mechanism of inhibition and find that the primary effect on conjugation is occlusion of the conjugative pilus by phage particles. This interaction is mediated primarily by phage coat protein g3p, and exogenous addition of the soluble fragment of g3p inhibited conjugation at low nanomolar concentrations. Our data are quantitatively consistent with a simple model in which association between the pili and phage particles or g3p prevents transmission of an F plasmid encoding tetracycline resistance. We also observe a decrease in the donor ability of infected cells, which is quantitatively consistent with a reduction in pili elaboration. Since many antibiotic-resistance factors confer susceptibility to phage infection through expression of conjugative pili (the receptor for filamentous phage), these results suggest that phage may be a source of soluble proteins that slow the spread of antibiotic resistance genes.

Show MeSH
Related in: MedlinePlus