Abstract
Environmental contamination has been viewed as an ecological malaise for which bioremediation can be prescribed as a “perfect medicine.” The solution to the problems with bioremediation lies in analyzing to what extent the microbes’ physiological machinery contributes to the degradation process and which biomolecules and their mechanisms are responsible for regulatory factors within the degradation system, such as protein, metabolite, and enzymatic chemical transformation. In the post-genomic era, recent advances in proteomics have allowed us to elucidate many complex biological mechanisms. Two-dimensional gel electrophoresis (2DE) in conjunction with mass spectrometry (MS) can be utilized to identify the biomolecules and their molecular mechanisms in bioremediation. A set of highly abundant global proteins over a pI range 4–7 was separated and compared by size fractionation (25–100 kDa) on 2DE. We identified a set of catabolic proteins, enzymes, and heat shock molecular chaperones associated with the regulatory network that was found to be overexpressed under phenol-stressed conditions. This chapter also offers optimized ideal directions for 2DE, followed by easy-to-follow directions for a protein identification strategy using MALDI-TOF and targeting novel proteins/enzymes for a universal set of experiments.
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References
Perera, F. P. (1997) Environment and cancer: who are susceptible? Science. 278, 1068–1073.
Chang, K. F., Fang, G. C., Chen, J. C., and Wu, Y. S. (2006) Atmospheric polycyclic aromatic hydrocarbons (PAHs) in Asia: a review from 1999 to 2004. Environ Pollut. 142, 388–396.
Boström, C. E., Gerde, P., Hanberg, A., Jernström, B., Johansson, C., Kyrklund, T., et al. (2002) Cancer risk assessment, indicators, and guidelines for polycyclic aromatic hydrocarbons in the ambient air. Environ. Health Perspect. 3, 451–488.
Belpomme, D., Irigaray, P., Hardell, L., Clapp, R., Montagnier, L., Epstein, S., et al. (2007) The multitude and diversity of environmental carcinogens. Environ. Res. 105, 414–429.
Peng, R. H., Xiong, A. S., Xue, Y., Fu, X. Y., Gao, F., Zhao, W., et al., (2008) Microbial biodegradation of polyaromatic hydrocarbons. FEMS Microbiol. Rev.(Epub ahead of print) PMID: 18662317.
Díaz, E. (2004) Bacterial degradation of aromatic pollutants: a paradigm of metabolic versatility. Int. Microbiol.7, 173–180.
Samanta, S. K., Singh, O. V., and Jain, R. K. (2002) Polycyclic aromatic hydrocarbons: environmental pollution and bioremediation. Trends Biotechnol. 20, 243–248.
Esteve-Nunez, A., Caballero, A., and Ramos, J.L. (2001) Biological degradation of 2,4,6-trinitrotoluene. Microbiol. Mol. Biol. Rev.65, 335–352.
Singh, O.V. (2007) Integrating “-omics” into biological processes and modeling for bioremediation. OMICS. 11, 231–232.
Zhao, B., and Poh, C. L. (2008) Insights into environmental bioremediation by microorganisms through functional genomics and proteomics. Proteomics.8, 874–881.
Singh, O. V. (2006) Proteomics and metabolomics: the molecular make-up of toxic aromatic pollutant bioremediation. Proteomics. 6, 5481–5492.
Wang, R. F., Wennerstrom, D., Cao, W. W., Khan, A. A., and Cerniglia, C. E. (2000) Cloning, expression, and characterization of the katG gene, encoding catalase-peroxidase, from the polycyclic aromatic hydrocarbon-degrading bacterium Mycobacterium sp. strain PYR-1. Appl. Environ. Microbiol. 66, 4300–4304.
Krivobok, S., Kuony, S., Meyer, C., Louwagie, M., Willison, J. C., and Jouanneau, Y. (2003) Identification of pyrene-induced proteins in Mycobacterium sp. strain 6PY1: evidence for two ring-hydroxylating dioxygenases. J. Bacteriol. 185, 3828–3841.
Kim, Y.H., Cho, K., Yun, S.H., Kim, J.Y., Kwon, K.H., Yoo J.S., et al. (2006) Analysis of aromatic catabolic pathways in Pseudomonas putida KT 2440 using a combined proteomics approach: 2DE/MS and cleavable isotope-coded affinity tag analysis. Proteomics. 6, 1301–1318.
Tomas-Gallardo, L., Canosa, I., Santero, E., Camafeita, E., Calvo, E., López, J. A., et al. (2006) Proteomic and transcriptional characterization of aromatic degradation pathways in Rhodoccocus sp. strain TFB. Proteomics. 6, S119–S132.
Santos, P. M., Roma, V., Benndorf, D., von Bergen, M., Harms, H., and Sá-Correia, I. (2007) Mechanistic insights into the global response to phenol in the phenol-biodegrading strain Pseudomonas sp. M1 revealed by quantitative proteomics. OMICS. 11, 233–251.
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Technical support rendered by Ms. Rashmi Singh for preparing this manuscript is gratefully acknowledged.
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Singh, O.V., Chandel, A.K. (2010). Two-Dimensional Gel Electrophoresis: Discovering Biomolecules for Environmental Bioremediation. In: Cummings, S. (eds) Bioremediation. Methods in Molecular Biology, vol 599. Humana Press. https://doi.org/10.1007/978-1-60761-439-5_10
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DOI: https://doi.org/10.1007/978-1-60761-439-5_10
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