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Isophthalate:coenzyme A ligase initiates anaerobic degradation of xenobiotic isophthalate

Isophthalate:coenzyme A ligase initiates anaerobic degradation of xenobiotic isophthalate

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JUNGHARE, Madan, Jasmin FREY, Khalid M. NAJI, Dieter SPITELLER, Gustav VAAJE-KOLSTAD, Bernhard SCHINK, 2022. Isophthalate:coenzyme A ligase initiates anaerobic degradation of xenobiotic isophthalate. In: BMC microbiology. BioMed Central. 22(1), 227. eISSN 1471-2180. Available under: doi: 10.1186/s12866-022-02630-x

@article{Junghare2022-09-28Isoph-58740, title={Isophthalate:coenzyme A ligase initiates anaerobic degradation of xenobiotic isophthalate}, year={2022}, doi={10.1186/s12866-022-02630-x}, number={1}, volume={22}, journal={BMC microbiology}, author={Junghare, Madan and Frey, Jasmin and Naji, Khalid M. and Spiteller, Dieter and Vaaje-Kolstad, Gustav and Schink, Bernhard}, note={Article Number: 227} }

Naji, Khalid M. Schink, Bernhard Naji, Khalid M. 2022-10-06T09:01:32Z Vaaje-Kolstad, Gustav Background: Environmental contamination from synthetic plastics and their additives is a widespread problem. Phthalate esters are a class of refractory synthetic organic compounds which are widely used in plastics, coatings, and for several industrial applications such as packaging, pharmaceuticals, and/or paints. They are released into the environment during production, use and disposal, and some of them are potential mutagens and carcinogens. Isophthalate (1,3-benzenedicarboxylic acid) is a synthetic chemical that is globally produced at a million-ton scale for industrial applications and is considered a priority pollutant. Here we describe the biochemical characterization of an enzyme involved in anaerobic degradation of isophthalate by the syntrophically fermenting bacterium Syntrophorhabdus aromaticivorans strain UI that activate isophthalate to isophthalyl-CoA followed by its decarboxylation to benzoyl-CoA.<br /><br />Results: Isophthalate:Coenzyme A ligase (IPCL, AMP-forming) that activates isophthalate to isophthalyl-CoA was heterologously expressed in E. coli (49.6 kDa) for biochemical characterization. IPCL is homologous to phenylacetate- CoA ligase that belongs to the family of ligases that form carbon-sulfur bonds. In the presence of coenzyme A, Mg2+ and ATP, IPCL converts isophthalate to isophthalyl-CoA, AMP and pyrophosphate (PPi). The enzyme was specifically induced after anaerobic growth of S. aromaticivorans in a medium containing isophthalate as the sole carbon source. Therefore, IPCL exhibited high substrate specificity and affinity towards isophthalate. Only substrates that are structurally related to isophthalate, such as glutarate and 3-hydroxybenzoate, could be partially converted to the respective coenzyme A esters. Notably, no activity could be measured with substrates such as phthalate, terephthalate and benzoate. Acetyl-CoA or succinyl-CoA did not serve as CoA donors. The enzyme has a theoretical pI of 6.8 and exhibited optimal activity between pH 7.0 to 7.5. The optimal temperature was between 25 °C and 37 °C. Denaturation temperature (Tm) of IPCL was found to be at about 63 °C. The apparent KM values for isophthalate, CoA, and ATP were 409 μM, 642 μM, and 3580 μM, respectively. Although S. aromaticivorans is a strictly anaerobic bacterium, the enzyme was found to be oxygen-insensitive and catalysed isophthalyl-CoA formation under both anoxic and oxic conditions.<br /><br />Conclusion: We have successfully cloned the ipcl gene, expressed and characterized the corresponding IPCL enzyme, which plays a key role in isophthalate activation that initiates its activation and further degradation by S. aromaticivorans. Its biochemical characterization represents an important step in the elucidation of the complete degradation pathway of isophthalate. 2022-10-06T09:01:32Z Isophthalate:coenzyme A ligase initiates anaerobic degradation of xenobiotic isophthalate Junghare, Madan eng Spiteller, Dieter Schink, Bernhard Junghare, Madan Spiteller, Dieter 2022-09-28 Attribution 4.0 International Vaaje-Kolstad, Gustav Frey, Jasmin Frey, Jasmin

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