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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

Spectrophotometric analysis for the heat-induced aggregation of MDH in the presence of AuNPs and Hsp70.1 μM MDH was suspended in assay buffer in the absence or mixed with high concentrations of AuNPs (0–100 μgmL-1) (a); the assay was repeated in the presence of 1.3 μM Hsp70 (b). The suspensions were subjected to heat stress at 48°C for 20 minutes. Absorbance values were measured at 340 nm in triplicates using a 96-well micro titre plate. Data are presented as mean and standard deviations.
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pone.0121243.g007: Spectrophotometric analysis for the heat-induced aggregation of MDH in the presence of AuNPs and Hsp70.1 μM MDH was suspended in assay buffer in the absence or mixed with high concentrations of AuNPs (0–100 μgmL-1) (a); the assay was repeated in the presence of 1.3 μM Hsp70 (b). The suspensions were subjected to heat stress at 48°C for 20 minutes. Absorbance values were measured at 340 nm in triplicates using a 96-well micro titre plate. Data are presented as mean and standard deviations.

Mentions: To further validate the findings, we monitored aggregation of MDH by taking spectrophotometric readings at 340 nm. The heat-induced aggregation of MDH results in turbidity which could be monitored by taking absorbance readings using a spectrophometer set at A340. As expected the Hsp70 chaperone suppressed the aggregation of MDH (Fig. 7). Furthermore, low levels of AuNPs (2.5–10 μgmL-1) suppressed the heat-induced aggregation of MDH in a concentration dependent fashion. In agreement with the findings obtained using SDS-PAGE analysis, higher levels (25–100 μgmL-1) of the AuNPs were not effective in suppressing the aggregation of MDH (Figs. 5B and 7A). The assay was repeated in the presence of 1.3 μM Hsp70. Hsp70 and the AuNPs at lower levels (2.5–10 μgmL-1) complemented each other in suppressing MDH aggregation (Fig. 7B). However, the aggregation of MDH in the presence of higher levels of AuNPs (25–100 μgmL-1) could not be suppressed by adding 1.3 μM Hsp70 to the reaction mix (Fig. 7B). This suggests that the citrate-coated AuNPs may have agglomerated at higher concentrations, presenting a surface curvature that compromised folding of MDH.


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

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

Spectrophotometric analysis for the heat-induced aggregation of MDH in the presence of AuNPs and Hsp70.1 μM MDH was suspended in assay buffer in the absence or mixed with high concentrations of AuNPs (0–100 μgmL-1) (a); the assay was repeated in the presence of 1.3 μM Hsp70 (b). The suspensions were subjected to heat stress at 48°C for 20 minutes. Absorbance values were measured at 340 nm in triplicates using a 96-well micro titre plate. Data are presented as mean and standard deviations.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0121243.g007: Spectrophotometric analysis for the heat-induced aggregation of MDH in the presence of AuNPs and Hsp70.1 μM MDH was suspended in assay buffer in the absence or mixed with high concentrations of AuNPs (0–100 μgmL-1) (a); the assay was repeated in the presence of 1.3 μM Hsp70 (b). The suspensions were subjected to heat stress at 48°C for 20 minutes. Absorbance values were measured at 340 nm in triplicates using a 96-well micro titre plate. Data are presented as mean and standard deviations.
Mentions: To further validate the findings, we monitored aggregation of MDH by taking spectrophotometric readings at 340 nm. The heat-induced aggregation of MDH results in turbidity which could be monitored by taking absorbance readings using a spectrophometer set at A340. As expected the Hsp70 chaperone suppressed the aggregation of MDH (Fig. 7). Furthermore, low levels of AuNPs (2.5–10 μgmL-1) suppressed the heat-induced aggregation of MDH in a concentration dependent fashion. In agreement with the findings obtained using SDS-PAGE analysis, higher levels (25–100 μgmL-1) of the AuNPs were not effective in suppressing the aggregation of MDH (Figs. 5B and 7A). The assay was repeated in the presence of 1.3 μM Hsp70. Hsp70 and the AuNPs at lower levels (2.5–10 μgmL-1) complemented each other in suppressing MDH aggregation (Fig. 7B). However, the aggregation of MDH in the presence of higher levels of AuNPs (25–100 μgmL-1) could not be suppressed by adding 1.3 μM Hsp70 to the reaction mix (Fig. 7B). This suggests that the citrate-coated AuNPs may have agglomerated at higher concentrations, presenting a surface curvature that compromised folding of MDH.

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