Many toxic compounds produced by human activities cannot be degraded by microorganisms available in natural water bodies and soil sediments.
But genetic engineering is being applied to generate microorganisms that may be capable of degrading them.
Some examples of such toxic compounds are: polychlorinated biphenyls (PCBs), chlorinated benzenes, and chlorinated phenols. Recently two strategies have been applied to the development of novel degradative pathways.
The first strategy, termed as pathway restructuring, is used when the resistant compound is similar in structure to a chemical compound that is degradable. It consists of determining which engyme-catalyzed steps of the known degradation pathway are unable to degrade these related compounds and therefore an attempt is made to modify the enzymes.
The second strategy, termed pathway assembly, is used when it is possible to plan a pathway that would utilize enzymes from several microorganisms and by means of genetic engineering those enzymes are assembled within a single host or in a consortium and placed under the appropriate metabolic regulation.
Pollution control by genetic engineering works best when pollutants are a known mixture of relatively concentrated organic compounds that are related to each other in structure, when conventional alternative organic nutrients are absent or perhaps added in controlled amounts only, and when there is no competition from indigenous microorganisms.
Thus, genetic engineering makes it possible to alter the properties of existing degradative enzymes, to modify regulatory mechanisms, and to assemble within single organism degradative enzymes from various different or phylogenetically distant organisms. However, the task of engineering an organism for the degradation of a particular target compound is not simple.
The easiest way to create an appropriate genetically engineered strain is to begin with an organism that already possesses much of the necessary degradative enzymatic machinery. An example of horizontal expansion of a catabolic pathway by genetic engineering is the following:
Psuedomonas putida carrying the TOL plasmid is able to grow on a variety of alkylbenzoates such as 3- and 4- methylbenzoate (3MB and 4MB), 3,4 dimethylbenzoate (3,4 DMB) and 3- ethylbenzoate (3-EB). However, the organism cannot grow on the very closely related compound 4- ethylbenzoate (4EB). To redesign the pathway to enable it to degrade 4 EB, it was necessary to broaden the effector specificity of the xylS regulator protein in a way that it would be activated by 4 EB, and to generate catechol 2,3- dioxygenase mutant enzymes capable of degrading 4- ethyl catechol. A genetic engineering strategy was applied for selection of mutants whose xylS protein would form a complex with 4 EB that could bind at Pmeta promoter and allow the transcription of genes controlled by this promoter (Ramos and Timmis, 1987).