Chaudhry 9316416 Carbamates and benzofurans are toxic pesticides that may be slowly biodegraded by microorganisms. This occurs as a hydrolytic process in some cells and as an oxidative process in others, and sometimes results in the release of more toxic intermediates than the pesticides themselves. We have chosen to study degradation of carbofuran (2,3-dihydro-2,2-dimethyl-7-benzofuranyl methylcarbamate) as a model compound to investigate the biochemical pathways and genetic determinants involved in the microbial degradation process. In previous studies several microorganisms were isolated that can utilize carbofuran as sole source of carbon or nitrogen. One of these, Pseudomonas sp. 50432, has the unique ability to metabolize carbamate by utilizing both hydrolytic and oxidative degradation pathways. Carbamates are hydrolyzed by a constitutively synthesized enzyme, while oxidation of the compound occurres only after the induction of the oxidative pathway. In the present research plan, we will: 1) isolate and purify the hydrolase involved in carbofuran degradation; 2) perform biochemical and molecular comparison of the Pseudomonas sp. 50432 hydrolase with enzymes in other microorganisms; and 3) clone the carbofuran hydrolase gene. The results of this study are expected to improve our understanding of the microbial catabolism of carbofuran and other carbamates. In addition, these findings should provide the basis for developing microbial agents capable of decontaminating soil and water polluted with carbamates, as well as in devising strategies for improving the efficacy and environmental safety of this group of pesticides. %%% Use of pesticides is vital for agricultural productivity. However, some pesticides, in particular the carbamate pesticides, are also toxic to animals and, although they may be slowly removed from the biosphere by microbes, this process is slow. The pesticides are now being found polluting ground water. In our laboratory, we will study how a microbe, which we named Pseudomonas sp 50432, can degrade carbofuran, one of the toxic polluting pesticides. To do this: 1) we will isolate and purify a key enzyme, the hydrolase, involved in degrading carbofuran; 2) we will determine if this enzyme is unique, or similar to enzymes in other related microbes; and 3) finally, we will clone and sequence of hydrolase gene. The results of this study are expected to improve our understanding of microbial destruction of carbofuran and other carbamate pesticides. In addition, these findings should provide the basis for developing microbial agents capable of decontaminating soil and water polluted with carbamate pesticides, as well as in devising strategies for improving the efficacy and the environmental safety of this group of pesticides. ***
