Two-dimensional gel electrophoresis: discovering biomolecules for environmental bioremediation.
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.
Singh
Om V
OV
Division of Biological and Health Science, University of Pittsburgh-Bradford, Bradford, PA 16701, USA. ovs11@pitt.edu
Chandel
Anuj K
AK
D001673:Biodegradation, Environmental; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D004783:Environmental Microbiology; D013058:Mass Spectrometry; D040901:Proteomics / Q000379:methods