In the beginning, the waters of the deep consisted of saline - a dilutesolution of sodium chloride. That chemical contained one atom each ofsodium and chlorine. The fossil counterpart of this ancient seawaterflows today in the veins of mammals, making blood salty.Down through the eons, more and more chlorine atoms came togetherin fluids more complex than seawater. Petroleum, for example, anatural product, implicates halogens. Soil bacteria over the ageslearned through mutation to degrade some of these compounds, thoughslowly.Then, as Homo sapiens industrialized, they created unnatural,xenobiotic, products - the human-made (or rather, anthropogenic)halogenated hydrocarbons. These ubiquitous solvents contain multiplechlorine atoms, which baffle the metabolic break-down capacities ofmicroorganisms. They tend to persist in the environment, and causeconcern for human health.Thus, the industrialized world is plagued with a myriad of apparentlyeternal toxic waste sites.Diverse industries use these compounds as solvents, degreasers andmachine-cleaners. Eastman Kodak dissolves its film emulsions indichloromethane, coats the film, then evaporates the halogenatedchemical. Computer manufacturers employ trichloroethylene to washoil off their chips.Bioremediation companies are busy trying to educate indigenous soilbacteria to degrade these indestructible residues. One of the mainproblems they face is a trap set by evolution itself.Early microorganisms grew up in an anaerobic, oxygen-free,environment, and learned how to metabolize one or two chlorine atomsfrom a given hydrocarbon. Later, aerobic organisms took over to finishthe job of stripping off halogens. Not many bacteria have been exposedto compounds with four of five chlorines, so they haven't evolved thegenes and enzymes to handle them.But the Environmental Protection Agency, which has declared war onchlorine contamination, can't wait millions of years for thesemicroorganisms to evolve. A University of Minnesota biochemist hascome up with a plan to speed up bacterial evolution.Lawrence Wackett's strategy for combining the anaerobic and aerobicbreak-down pathways in a single microorganism appeared inyesterday's (April 14) Nature. Its title: "Metabolism ofpolyhalogenated compounds by a genetically engineered bacterium."That smartened-up microorganism is recombinant Pseudomonasputida, which deploys seven genes encoding two enzyme systems.These deliver a one-two anaerobic-aerobic punch to the likes ofpentachloroethane - CCl3CHCl2. This solvent contains five chlorineatoms. By the time recombinant P. putida is finished tearing it apart,the highly halogenated chemical has been broken down, first totrichlorethylene, then to manageable metabolites, and its chlorinesimmobilized as simple chlorides.Wackett, who describes his scientific discipline as ecological proteinbiochemistry, teaches at the University of Minnesota. His key concept,he explained to BioWorld Today, was "to take two existing enzymesystems known to catalyze other reactions, and construct amicroorganism in which the first would partially dechlorinate somehighly chlorinated compounds, then becomes a substrate for thesecond, which then completes the process."This tandem harnessing of two enzyme systems in a single bacterium,Wackett said, "was a new twist, which perhaps is why Nature found itappealing."In his construct, the first multi-component enzyme, cytochromeP450cam, metabolizes camphor, a natural plant product, and deploysthree genes in the process. The second is toluene dioxygenase; its four-gene enzyme complex degrades toluene, which is also a naturalproduct, found in crude oil."They've probably been around in the environment for millions ofyears," Wackett observed.He and his colleagues sicced their P. putida onto five-chlorinepentachlorethane, as their model waste-product target.Wackett's "new twist" aims to overcome the "cumbersome andexpensive transfer of material" from an initial anaerobic fermenter to afinal aerobic bioreactor. "That's part of our idea too," he said. "If wecould get a single microorganism to do both parts of the process in onepot, ultimately, in terms of engineering, it would make the system workcheaply."Eventually, the Minnesota team aims to try the system in pilot-scalereactors, and thereafter in field conditions. But Wackett's next step is tostrengthen P. putida's genetic stability by moving the seven genes fromexpression plasmids into the organism's chromosome.In nature, those genes are scattered among existing strains of thebacterium, which already has a reputation for degrading organiccompounds such as aromatic hydrocarbons and some crude-oilcomponents, but not heavily chlorinated or fluorinated compounds.Wackett's study also metabolized chlorfluorocarbons, which, he pointsout, "are implicated in ozone-layer destruction."Peter Chapman, a senior microbiologist at the EPA's Gulf BreezeEnvironmental Research Laboratory, had this to say to BioWorldToday concerning Wackett's work, which he knows well:"I think that first and foremost it points in directions that are valuable,without necessarily presenting us with an organism that is immediatelygoing to be used in the technology. It provides us with the knowledgethat constructing organisms to degrade xenobiotics is somethingeminently feasible. From a knowledge of enzyme reaction mechanisms,one could rationally design such organisms."
-- David N. Leff Science Editor
(c) 1997 American Health Consultants. All rights reserved.