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Chapter 30 : Microbial Bioremediation of Chemical Pollutants: How Bacteria Cope with Multi-Stress Environmental Scenarios
Category: Microbial Genetics and Molecular Biology
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This chapter discusses the aspects of bioremediation that are related to metabolism of recalcitrant chemicals by bacteria, leaving out also detoxification and immobilization of metal ions and metalloids. The most frequent types of sites amenable to bioremediation include soil, freshwater, seawater, and sediments. The chapter talks about the instances where recalcitrant and/or xenobiotic compounds are endowed with chemical properties that cause a deleterious effect on the catalytic microorganisms present in the site-regardless of whether they can be ultimately metabolized. The chemicals at stake include metals, chaotropic agents, aromatics, and hydrophobic compounds. These stressors can be grouped based on their effect on bacterial metabolism. Heat shock-like stress and oxidative damage are certainly the two more prevalent conditions endured by environmental bacteria during in situ biodegradation of chemical waste. This is true for singular stressors as well as for mixtures of them, the most frequent bioremediation scenarios. Setting up suitable experimental systems to examine this issue is an important point in the current bioremediation research agenda. The bottleneck for the successful catabolism of a recalcitrant compound is most often not the nature of the biochemical route for its degradation, but the overcoming of the endogenous and exogenous stress associated to the working conditions.
Organization of the TOL operons of plasmid pWW0. The plasmid encodes a complete enzymatic machinery for the stepwise oxidation of one of the methyl groups of toluene, m- xylene and p- xylene (but not o- xylene), into the corresponding carboxylic acids (encoded by the so-called upper operon, driven by the σ54 Pu promoter). This is followed by ring dioxygenation and meta- cleavage of the resulting (methyl)catechol all the way down to the TCA (encoded by the meta or lower pathway, driven by the Pm promoter). Expression of each of the gene clusters is subject to the interplay between host factors and plasmid-encoded regulators. The later include XylR (an m- xylene-responsive σ54-dependent activator acting on Pu and Ps) and XylS (an m- toluate-responsive factor acting on Pm). Activation of the meta operon requires also the concourse of either the heat shock σ32 or the stationary stress σ38 of the host (Dominguez-Cuevas et al., 2005 ; Marques et al., 1999 ). Overproduction of XylS can also activate Pm in the absence of cognate effectors. XylR expression is autoregulated.
The economy of stress versus biodegradation in environmental bacteria. (a) The Gestalt (perception) chalice. This cartoon represents the notorious fact, known to the Gestalt branch of cognitive science, that it is often impossible to simultaneously perceive two different aspects of the same thing: either the faces or the chalice, but not both. This image is used here as a metaphor for the dual and altogether different notice that environmental microbes take of the presence of recalcitrant chemicals, as stressors as well as nutrients. (b) Regulatory choices in environmental bacteria. The accessible energy and reducing power in single cells is operatively represented here as ATP and NAD(P)H. Bacteria must spend a considerable share of such energy for the buildup of catabolic pathways for the dedicated metabolism of available C-sources (and to a lesser extent N, P sources and oligonutrients as well). Yet, coping with both external stress (left) or the toxicity caused by some of the C sources (right) consumes a share of the energy/reduction equivalents produced. (c) Only if the balance is positive, can biodegradation occur, as sketched.
Biocatalysis under endogenous and exogenous environmental stress. One key difference between chemical catalysis in the laboratory and its counterparts during bioremediation is the large number of factors at play in the process. There is first the issue of bioavailability because the substrate of the desired reaction (A) may not directly access the core catalyst (often an intracellular enzyme) because of its intrinsic lack of solubility or its adsorption to mineral matrices. In addition, the substrate is often a stressor for the live bacterial catalysts. Furthermore, the desired reactions may not be coupled to growth and can originate side products (C, F) with deleterious effects on the microorganism. Finally, the product of the process (Z) might be toxic and inhibit the progression of the whole biotransformation. The external milieu also brings in exogenous stresses that affect the course of action. This scenario provides an optimal chance to apply a systems biology approach to the design of processes of this sort (de Lorenzo, 2008 ).
Control of the stress versus metabolism tradeoff in σ54 promoters. Because σ54 promoters, such as Pu of the TOL plasmid, can be transcribed in vitro by just combining purified integration host factor, the sigma factor core RNAP, and activated XylR, it is plausible that the mechanism(s) in vivo that adapt transcription to environmental stresses do so by regulating the binding of one of more of these proteins to their target sites. In addition, the upstream activating sequences (UAS) of Pu appear to be mostly unoccupied at all growth stages. This gives rise to multiple regulatory possibilities. In cases where the promoter has little intrinsic affinity for the σ54 -RNAP, occupation of the -12/-24 region depends exclusively on the successful competition of sigmas at the stationary phase, governed by ppGpp. In this case, physiological control mostly reflects such competition. Some sigmas, like σ70, can themselves be checked by other anti-sigma proteins such as Rsd (Yuan et al., 2008 ). In the other extreme, σ54 promoters with a high intrinsic affinity for the enzyme may have the -12/-24 motif occupied even at the low concentrations of σ54 -RNAP available prior to stationary phase. In these instances, the promoter may not be subject to any physiological or stress-related inhibition, but may be active throughout all growth states. A number of extra factors (TurA, PprA) control the binding of core transcription components to their target sequences (Rescalli et al., 2004 ; Vitale et al., 2008 ).