Heterologous Expression of the Phytochelatin Synthase CaPCS2 from Chlamydomonas acidophila and Its Effect on Different Stress Factors in Escherichia coli
Abstract
:1. Introduction
2. Materials and Methods
2.1. Bacterial Strains and Growth Conditions
2.2. Analysis of Exposure to Metal(loid)s, Salt, and Acid pH by Growth Curve Tests
2.3. Test for Resistance to UVB Radiation and Exposure to Perchlorate Using the Droplet Test
2.4. Determination of Cellular Metal Concentration Using Inductively Coupled Plasma Spectroscopy-Mass Spectrometry (ICP-MS) Quantitative Analysis
2.5. Statistics
2.6. Gene Characterization
2.7. Phylogenetic Analysis
3. Results
3.1. Tolerance to Metal(loid) Exposure
3.2. Tolerance to UVB Radiation and Exposure to Perchlorate by Droplet Test
3.3. Tolerance to Acidic pH and Salt Exposure
3.4. Determination of Cellular Metal Concentration
3.5. CaPCS2 Gene from Chlamydomonas Acidophila
4. Discussion
4.1. Role of Phytochelatin Sythases in Tolerance to Abiotic Stressors
4.2. Phytochelatin Sythetase and Heavy Metal(loid)s Bioaccumulation
5. Conclusions
- Heterologous expression of the PCS gene from C. acidophila (CaPCS2) confers resistance in E. coli to several metal(loid)s that directly or indirectly produce ROS. This PCS is not involved in the bioaccumulation of metal(loid)s in the cell.
- The E. coli strain transformed with the CaPCS2 gene showed increased resistance to exposure to various cellular stress factors, such as the presence of perchlorate and exposure to UVB radiation. This strain also exhibited greater tolerance to the presence of salt and acidic pH in the culture medium than the control strain.
- The phylogenetic analysis of the CaPCS2 gene showed characteristics intermediate to eukaryotic and prokaryotic PCS genes. The amino acid sequence suggests closer relationships to bacterial PCS than to plant PCS. However, the complete gene sequence contains introns, such as eukaryotic genes.
- The strictly conserved Cys residues and catalytic triad (Cys56, His162, and Asp180) are also present in CaPCS2 (Cys70, His189, and Asp207). CaPCS2 lacks four of the five conserved Cys residues at the N-terminus, such as bacterial PCS.
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Metal/ | Concentration (mM) | Bioaccumulation | Bioaccumulation | t-Test | ||
---|---|---|---|---|---|---|
Metalloid | (mgg−1d.w) | (mgg−1d.w) | For Equality ofMeans | |||
NC | SD | CaPCS2 | SD | Sig(2-tailed) | ||
1 | 6299 | 1814 | 6624 | 2734 | 0.902 | |
Cu | 2 | 1006 | 116 | 1111 | 59 | 0.232 |
4 | 3822 | 1192 | 4204 | 1536 | 0.504 | |
Cd | 0.3 | 4653 | 893 | 4640 | 1294 | 0.982 |
1 | 13,377 | 3461 | 14,547 | 2773 | 0.533 | |
As | 1 | 246 | 63 | 272 | 70 | 0.513 |
0.5 | 381 | 25 | 338 | 148 | 0.642 | |
Ni | 1 | 1375 | 102 | 1017 | 200 | 0.051 |
4 | 8486 | 1099 | 6234 | 1365 | 0.011 * |
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Díaz, S.; Aguilera, Á.; de Figueras, C.G.; de Francisco, P.; Olsson, S.; Puente-Sánchez, F.; González-Pastor, J.E. Heterologous Expression of the Phytochelatin Synthase CaPCS2 from Chlamydomonas acidophila and Its Effect on Different Stress Factors in Escherichia coli. Int. J. Environ. Res. Public Health 2022, 19, 7692. https://doi.org/10.3390/ijerph19137692
Díaz S, Aguilera Á, de Figueras CG, de Francisco P, Olsson S, Puente-Sánchez F, González-Pastor JE. Heterologous Expression of the Phytochelatin Synthase CaPCS2 from Chlamydomonas acidophila and Its Effect on Different Stress Factors in Escherichia coli. International Journal of Environmental Research and Public Health. 2022; 19(13):7692. https://doi.org/10.3390/ijerph19137692
Chicago/Turabian StyleDíaz, Silvia, Ángeles Aguilera, Carolina G. de Figueras, Patricia de Francisco, Sanna Olsson, Fernando Puente-Sánchez, and José Eduardo González-Pastor. 2022. "Heterologous Expression of the Phytochelatin Synthase CaPCS2 from Chlamydomonas acidophila and Its Effect on Different Stress Factors in Escherichia coli" International Journal of Environmental Research and Public Health 19, no. 13: 7692. https://doi.org/10.3390/ijerph19137692