Bioremediation of catecholic pollutants with novel oxygen-insensitive catechol 2,3-dioxygenase and its potential in biomonitoring of catechol in wastewater.

The oxygenases are essential in the bioremediation of xenobiotic pollutants. To overcome cultivability constraints, this study aims to identify new potential extradiol dioxygenases using the functional metagenomics approach. RW1-4CC, a novel catechol 2,3-dioxygenase, was isolated using functional metagenomics approach, expressed in a heterologous system, and characterized thoroughly using state-of-the-art techniques. The serial truncation mutations of the C-terminal tail increase the catalytic efficiency of truncated proteins against the 2,3-dihydroxybiphenyl (2,3-DHB). RW1-4CC lose its 50% of activity at 60 °C, with its optimum temperature at 15 °C, whereas the truncated proteins were found to be more stable at extended temperature range, i.e., both RW1-4CC-A and RW1-4CC-B retained 50% of their activity at 75 °C, with their temperature optima at 55 °C and 65 °C, respectively. The molecular docking studies further confirmed the high binding affinity of truncated proteins for the 2,3-DHB than catechol. The molecular modeling analysis revealed the difference in iron-binding and substrate interacting environment of RW1-4CC and its truncated proteins. The efficiency of purified RW1-4CC to detect catechol was evaluated using a gold screen-printed electrode by cyclic voltammetry. RW1-4CC detected catechol in wastewater and artificial seawater up to the concentration of 100 μm, which makes it reliable for catechol detection.