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Protective role of bacillithiol in superoxide stress and Fe-S metabolism in Bacillus subtilis.

Fang Z, Dos Santos PC - Microbiologyopen (2015)

Bottom Line: Interestingly, Fe-S cluster containing isopropylmalate isomerase (LeuCD) and glutamate synthase (GOGAT) showed decreased activities in BSH strain.Deficiency of BSH also resulted in decreased levels of intracellular Fe accompanied by increased levels of manganese and altered expression levels of Fe-S cluster biosynthetic SUF components.Together, this study is the first to establish a link between BSH and Fe-S metabolism in B. subtilis.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Wake Forest University, Winston-Salem, North Carolina, 27016.

No MeSH data available.


Related in: MedlinePlus

Specific activities of ACN (A) and GOGAT (B) under stress conditions. Cells were grown in MM to late log phase (OD600 of 0.8–1.0 measured with a 1 cm cuvette), at 37°C, and then 100 μmol/L of 2,2′-dypridyl (DP), 100 μmol/L of paraquat (PQ), 100 μmol/L of H2O2 (HP), 50 μmol/L of CdCl2 (Cd), or 100 μmol/L of CuCl (Cu) was added to challenge the cultures for 30 min. Cell lysates were prepared as described in Experimental Procedures. The wild-type strain is represented in black bars, whereas ΔbshA strain is shown in gray bars. All assays were repeated in triplicates. The statistical analysis was performed using unpaired t test, P values compare ΔbshA sample with wild-type sample (NS, not significant, *P < 0.05, **P < 0.01, ***P < 0.001).
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fig04: Specific activities of ACN (A) and GOGAT (B) under stress conditions. Cells were grown in MM to late log phase (OD600 of 0.8–1.0 measured with a 1 cm cuvette), at 37°C, and then 100 μmol/L of 2,2′-dypridyl (DP), 100 μmol/L of paraquat (PQ), 100 μmol/L of H2O2 (HP), 50 μmol/L of CdCl2 (Cd), or 100 μmol/L of CuCl (Cu) was added to challenge the cultures for 30 min. Cell lysates were prepared as described in Experimental Procedures. The wild-type strain is represented in black bars, whereas ΔbshA strain is shown in gray bars. All assays were repeated in triplicates. The statistical analysis was performed using unpaired t test, P values compare ΔbshA sample with wild-type sample (NS, not significant, *P < 0.05, **P < 0.01, ***P < 0.001).

Mentions: Cu(I) can strongly trigger Fenton-like reactions to produce reactive hydroxyl radicals, which further cause a global oxidative stress response, such as decreased activities for Fe–S enzymes (Macomber and Imlay 2009). As shown in Figure3A, the addition of Cu(I) did not inhibit the initial growth rate of the wild-type strain, but decreased the OD600 in stationary phase from 0.4 to 0.265, displaying similar growth inhibition as previously reported for B. subtilis (Chillappagari et al. 2010). The BSH mutant, however, showed a concentration-dependent inhibition of the initial growth rate, which indicated the protective role of BSH against Cu(I) stress. The addition of Cu(I) slightly decreased the concentration of BSH (from 0.68 to 0.44 μmol/g DW) and BSSB (from 0.052 to 0.035 μmol/g DW), but it significantly stimulated protein S-bacillithiolation by sixfold (from 0.02 to 0.12 μmol/g DW) (Fig.5A). Wild-type cells when cultured in MM and challenged with Cu(I) for 30 min showed reduction of 50% and 87% of ACN and GOGAT activities (Fig.4). This supported previous proposals that Fe–S enzymes were the targets for Cu(I) stress in E. coli (Macomber and Imlay 2009) and B. subtilis (Macomber and Imlay 2009; Chillappagari et al. 2010). However, in the mutant strain the activities of these enzymes were not further affected with Cu(I) challenge (Fig.4).


Protective role of bacillithiol in superoxide stress and Fe-S metabolism in Bacillus subtilis.

Fang Z, Dos Santos PC - Microbiologyopen (2015)

Specific activities of ACN (A) and GOGAT (B) under stress conditions. Cells were grown in MM to late log phase (OD600 of 0.8–1.0 measured with a 1 cm cuvette), at 37°C, and then 100 μmol/L of 2,2′-dypridyl (DP), 100 μmol/L of paraquat (PQ), 100 μmol/L of H2O2 (HP), 50 μmol/L of CdCl2 (Cd), or 100 μmol/L of CuCl (Cu) was added to challenge the cultures for 30 min. Cell lysates were prepared as described in Experimental Procedures. The wild-type strain is represented in black bars, whereas ΔbshA strain is shown in gray bars. All assays were repeated in triplicates. The statistical analysis was performed using unpaired t test, P values compare ΔbshA sample with wild-type sample (NS, not significant, *P < 0.05, **P < 0.01, ***P < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4554457&req=5

fig04: Specific activities of ACN (A) and GOGAT (B) under stress conditions. Cells were grown in MM to late log phase (OD600 of 0.8–1.0 measured with a 1 cm cuvette), at 37°C, and then 100 μmol/L of 2,2′-dypridyl (DP), 100 μmol/L of paraquat (PQ), 100 μmol/L of H2O2 (HP), 50 μmol/L of CdCl2 (Cd), or 100 μmol/L of CuCl (Cu) was added to challenge the cultures for 30 min. Cell lysates were prepared as described in Experimental Procedures. The wild-type strain is represented in black bars, whereas ΔbshA strain is shown in gray bars. All assays were repeated in triplicates. The statistical analysis was performed using unpaired t test, P values compare ΔbshA sample with wild-type sample (NS, not significant, *P < 0.05, **P < 0.01, ***P < 0.001).
Mentions: Cu(I) can strongly trigger Fenton-like reactions to produce reactive hydroxyl radicals, which further cause a global oxidative stress response, such as decreased activities for Fe–S enzymes (Macomber and Imlay 2009). As shown in Figure3A, the addition of Cu(I) did not inhibit the initial growth rate of the wild-type strain, but decreased the OD600 in stationary phase from 0.4 to 0.265, displaying similar growth inhibition as previously reported for B. subtilis (Chillappagari et al. 2010). The BSH mutant, however, showed a concentration-dependent inhibition of the initial growth rate, which indicated the protective role of BSH against Cu(I) stress. The addition of Cu(I) slightly decreased the concentration of BSH (from 0.68 to 0.44 μmol/g DW) and BSSB (from 0.052 to 0.035 μmol/g DW), but it significantly stimulated protein S-bacillithiolation by sixfold (from 0.02 to 0.12 μmol/g DW) (Fig.5A). Wild-type cells when cultured in MM and challenged with Cu(I) for 30 min showed reduction of 50% and 87% of ACN and GOGAT activities (Fig.4). This supported previous proposals that Fe–S enzymes were the targets for Cu(I) stress in E. coli (Macomber and Imlay 2009) and B. subtilis (Macomber and Imlay 2009; Chillappagari et al. 2010). However, in the mutant strain the activities of these enzymes were not further affected with Cu(I) challenge (Fig.4).

Bottom Line: Interestingly, Fe-S cluster containing isopropylmalate isomerase (LeuCD) and glutamate synthase (GOGAT) showed decreased activities in BSH strain.Deficiency of BSH also resulted in decreased levels of intracellular Fe accompanied by increased levels of manganese and altered expression levels of Fe-S cluster biosynthetic SUF components.Together, this study is the first to establish a link between BSH and Fe-S metabolism in B. subtilis.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Wake Forest University, Winston-Salem, North Carolina, 27016.

No MeSH data available.


Related in: MedlinePlus