Identification and characterisation of moderately thermostable diisobutyl phthalate degrading esterase from a Great Artesian Basin Bacillus velezensis NP05

Phthalate esters are known to be endocrine disrupting chemicals and are documented to pollute environments. Enzymatic degradation of PAEs is a potential bioremedial strategy to manage contamination. Thermostable bioremedial enzymes have advantages in enzyme manufacturing and storage. In this study, we identified, overexpressed, and characterised a moderately thermostable para-nitrobenzyl esterase from whole genome sequencing of a Bacillus velezensis NP05 from the Great Artesian Basin, capable of sequential 2-step hydrolysis of diisobutyl phthalate. The pnbA enzyme has a molecular weight of 55.14 kDa and pI of 5.31. It preferentially degrades para-nitrophenyl butanoate and has an optimal pH of 7-8. The pnbA esterase has an optimal temperature of 55°C with a half-life of 4 hours. Using HPLC we found that pnbA (0.122 U) can hydrolyse 0.83mM of DIBP within 25 minutes. Lastly, pnbA is potentially a more economically viable candidate for enzymatic bioremediation of diisobutyl phthalate as a free enzyme.