Limits...
DnaK protein alleviates toxicity induced by citrate-coated gold nanoparticles in Escherichia coli.

Makumire S, Revaprasadu N, Shonhai A - PLoS ONE (2015)

Bottom Line: We further investigated the effects of the AuNPs on the solubility of the E. coli BB1553 proteome.The toxic effects of the AuNPs were alleviated by transforming the E. coli BB1553 cells with a construct expressing DnaK.Our study suggests a role for DnaK in alleviating nanoparticle induced stress in E. coli.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, School of Mathematics & Natural Sciences, University of Venda, Thohoyandou, South Africa; Department of Biochemistry & Microbiology, University of Zululand, KwaDlangezwa, South Africa.

ABSTRACT
A number of previously reported studies suggest that synthetic gold nanoparticles (AuNPs) are capable of stabilising proteins against heat stress in vitro. However, it remains to be understood if AuNPs confer stability to proteins against cellular stress in vivo. Heat shock proteins (Hsps) are conserved molecules whose main role is to facilitate folding of other proteins (chaperone function). Hsp70 (called DnaK in prokaryotes) is one of the most prominent molecular chaperones. Since gold nanoparticles exhibit chaperone-like function in vitro, we investigated the effect of citrate-coated gold nanoparticles on the growth of E. coli BB1553 cells that possess a deleted dnaK gene. We further investigated the effects of the AuNPs on the solubility of the E. coli BB1553 proteome. E. coli BB1553 cells exposed to AuNPs exhibited cellular defects such as filamentation and plasma membranes pulled off the cell wall. The toxic effects of the AuNPs were alleviated by transforming the E. coli BB1553 cells with a construct expressing DnaK. We also noted that cells in which DnaK was restored exhibited distinct zones to which the nanoparticles were restricted. Our study suggests a role for DnaK in alleviating nanoparticle induced stress in E. coli.

No MeSH data available.


Related in: MedlinePlus

Proposed model illustrating the effects of citrate-coated gold nanoparticles in vitro and in E. coli cells.The model describes the proposed effects of single species versus agglomerated species of citrate-gold nanoparticles on the integrity of proteins in vitro and in E. coli cells that are deficient of DnaK and in which DnaK function is restored.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4383610&req=5

pone.0121243.g008: Proposed model illustrating the effects of citrate-coated gold nanoparticles in vitro and in E. coli cells.The model describes the proposed effects of single species versus agglomerated species of citrate-gold nanoparticles on the integrity of proteins in vitro and in E. coli cells that are deficient of DnaK and in which DnaK function is restored.

Mentions: To further confirm the effect of citrate coated AuNPs on protein integrity, we exposed an aggregation prone protein, MDH, to heat stress in the absence and in the presence of variable levels of AuNPs (Figs. 5 and 6). We observed that at lower order levels (2.5–10 μgmL-1), AuNPs effectively suppressed heat-induced aggregation of MDH. However, at higher order levels (25–100 μgmL-1), AuNPs were not effective in suppressing the heat-induced aggregation of MDH (Figs. 5B and 7A-B). AuNPs at higher concentrations tend to agglomerate [11][38]. Under such conditions, they possibly present a surface curvature that is detrimental to protein stability. It is interesting to note that the addition of Hsp70 to MDH in the presence of high levels of AuNPs did not result in reduction of MDH aggregation (Fig. 7B). This suggests that at high levels, AuNPs may have promoted protein aggregation that Hsp70 failed to reverse in vitro. However, in the cell, Hsp70 is more effective in reversing aggregates as it cooperates with other molecular chaperones [35]. This could explain why the restoration of DnaK in E. coli BB1553 cells promoted cell recovery even though the internalised AuNPs appeared agglomerated. We hypothesize that the AuNP nucleation sites created a platform that promoted protein misfolding and aggregation (Fig. 8). Since DnaK has a propensity to bind to misfolded protein [14], it is possible it was recruited to the nucleation sites by misfolded proteins, leading to the formation of the distinctly dense structures viewed under TEM (Fig. 3, panels C2, D2; Fig. 8). Subsequently, DnaK, possibly in cooperation with other chaperones in the cell effectively reversed protein aggregation, leading to cell survival. In the absence of DnaK, we propose that the agglomerations formed by the AuNPs in the absence of DnaK promoted irreversible protein aggregation. This is supported by the observation that E. coli DnaK- cells exposed to AuNPs had a higher proportion of proteins in the pellet fraction, compared to cells that were not exposed to AuNPs. On the other hand, E. coli DnaK+ cells that were exposed to AuNPs exhibited similar protein solubility pattern to that observed in E. coli DnaK+ cells that were not exposed to AuNPs (Fig. 4).


DnaK protein alleviates toxicity induced by citrate-coated gold nanoparticles in Escherichia coli.

Makumire S, Revaprasadu N, Shonhai A - PLoS ONE (2015)

Proposed model illustrating the effects of citrate-coated gold nanoparticles in vitro and in E. coli cells.The model describes the proposed effects of single species versus agglomerated species of citrate-gold nanoparticles on the integrity of proteins in vitro and in E. coli cells that are deficient of DnaK and in which DnaK function is restored.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4383610&req=5

pone.0121243.g008: Proposed model illustrating the effects of citrate-coated gold nanoparticles in vitro and in E. coli cells.The model describes the proposed effects of single species versus agglomerated species of citrate-gold nanoparticles on the integrity of proteins in vitro and in E. coli cells that are deficient of DnaK and in which DnaK function is restored.
Mentions: To further confirm the effect of citrate coated AuNPs on protein integrity, we exposed an aggregation prone protein, MDH, to heat stress in the absence and in the presence of variable levels of AuNPs (Figs. 5 and 6). We observed that at lower order levels (2.5–10 μgmL-1), AuNPs effectively suppressed heat-induced aggregation of MDH. However, at higher order levels (25–100 μgmL-1), AuNPs were not effective in suppressing the heat-induced aggregation of MDH (Figs. 5B and 7A-B). AuNPs at higher concentrations tend to agglomerate [11][38]. Under such conditions, they possibly present a surface curvature that is detrimental to protein stability. It is interesting to note that the addition of Hsp70 to MDH in the presence of high levels of AuNPs did not result in reduction of MDH aggregation (Fig. 7B). This suggests that at high levels, AuNPs may have promoted protein aggregation that Hsp70 failed to reverse in vitro. However, in the cell, Hsp70 is more effective in reversing aggregates as it cooperates with other molecular chaperones [35]. This could explain why the restoration of DnaK in E. coli BB1553 cells promoted cell recovery even though the internalised AuNPs appeared agglomerated. We hypothesize that the AuNP nucleation sites created a platform that promoted protein misfolding and aggregation (Fig. 8). Since DnaK has a propensity to bind to misfolded protein [14], it is possible it was recruited to the nucleation sites by misfolded proteins, leading to the formation of the distinctly dense structures viewed under TEM (Fig. 3, panels C2, D2; Fig. 8). Subsequently, DnaK, possibly in cooperation with other chaperones in the cell effectively reversed protein aggregation, leading to cell survival. In the absence of DnaK, we propose that the agglomerations formed by the AuNPs in the absence of DnaK promoted irreversible protein aggregation. This is supported by the observation that E. coli DnaK- cells exposed to AuNPs had a higher proportion of proteins in the pellet fraction, compared to cells that were not exposed to AuNPs. On the other hand, E. coli DnaK+ cells that were exposed to AuNPs exhibited similar protein solubility pattern to that observed in E. coli DnaK+ cells that were not exposed to AuNPs (Fig. 4).

Bottom Line: We further investigated the effects of the AuNPs on the solubility of the E. coli BB1553 proteome.The toxic effects of the AuNPs were alleviated by transforming the E. coli BB1553 cells with a construct expressing DnaK.Our study suggests a role for DnaK in alleviating nanoparticle induced stress in E. coli.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, School of Mathematics & Natural Sciences, University of Venda, Thohoyandou, South Africa; Department of Biochemistry & Microbiology, University of Zululand, KwaDlangezwa, South Africa.

ABSTRACT
A number of previously reported studies suggest that synthetic gold nanoparticles (AuNPs) are capable of stabilising proteins against heat stress in vitro. However, it remains to be understood if AuNPs confer stability to proteins against cellular stress in vivo. Heat shock proteins (Hsps) are conserved molecules whose main role is to facilitate folding of other proteins (chaperone function). Hsp70 (called DnaK in prokaryotes) is one of the most prominent molecular chaperones. Since gold nanoparticles exhibit chaperone-like function in vitro, we investigated the effect of citrate-coated gold nanoparticles on the growth of E. coli BB1553 cells that possess a deleted dnaK gene. We further investigated the effects of the AuNPs on the solubility of the E. coli BB1553 proteome. E. coli BB1553 cells exposed to AuNPs exhibited cellular defects such as filamentation and plasma membranes pulled off the cell wall. The toxic effects of the AuNPs were alleviated by transforming the E. coli BB1553 cells with a construct expressing DnaK. We also noted that cells in which DnaK was restored exhibited distinct zones to which the nanoparticles were restricted. Our study suggests a role for DnaK in alleviating nanoparticle induced stress in E. coli.

No MeSH data available.


Related in: MedlinePlus