Bacillus licheniformis M2-7 is a heat-resistant bacterium able to biotransform polycyclic aromatic hydrocarbons. It can transform a wide range of these compounds as naphthalene, phenanthrene, pyrene and benzo[a]pyrene. Benzo[a]pyrene is a polycyclic aromatic hydrocarbon of high molecular weight considered as potentially toxic and carcinogenic for humans. Aiming to discover the genes involved in the biotransformation of benzo[a]pyrene, we made a B. licheniformis M2-7 genomic library in E. coli. We isolated two E. coli strains that were able to grow in minimal salt medium supplemented with benzo[a]pyrene. From the analysis of the DNA fragments in the clones H23 and H38, we identified open reading frames coding for 5 possible genes, among them pobA and fabHB, which products are the enzymes 4hydroxybenzoate 3-monooxygenase and the ketoacyl-ACP synthase Genetics and Molecular Research 17 (2): gmr16039916 III, respectively. To evaluate the role of these genes in the metabolism of benzo[a]pyrene in B. licheniformis M2-7, we estimated their relative expression through reverse transcription quantitative PCR. Finally, we observed that the genes pobA and fabHB were overexpressed after 3 h under induction with benzo[a]pyrene, suggesting that this strain could use these genes during the metabolism of this PAH, plus it does it in a faster time than that reported for other bacterial genera
Benzopyrene is a high molecular weight polycyclic aromatic hydrocarbon with highly recalcitrant and develops carcinogenic effects. CsrA is a conserved regulatory protein, which controls the translation and stability of its target transcripts, having negative or positive effects depending on the target mRNAs. It is known that Bacillus licheniformis M2-7 has the ability to grow and survive in concentrations of hydrocarbons as benzopyrene, which prompted in part by CsrA, as it occurs in the presence of gasoline.However, there are a few studies that reveal the genes involved in that process. In order to know the involved genes in the Bacillus licheniformis M2-7 degradation pathway, the plasmid pCAT-sp containing a mutation in the catEgen was constructed and used to transform B. licheniformis M2-7 and generate the CAT1 strain. We determined the capacity of the mutant B. licheniformis(CAT1) to grow in the presence of glucose and benzopyrene as a carbon source. We observed that CAT1 strain presented an increase in growth in the presence of glucose, but a statistically considerable decrease in the presence of benzopyrene with respect to the wild strain M2-7. Also we demonstrated that the Csr system regulates its expression positively since it was observed that the expression of the gene in the mutant strain LYA12 (M2-7 csrA:: Sp, SpR) decreased considerably with respect to the wild strain. This allowed to characterize the metabolic pathway that Bacillus licheniformis M2-7 implements in the presence of benzopyrene.
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