Enhanced biodegradation of mixed PAHs by mutated naphthalene 1,2-dioxygenase encoded by Pseudomonas putida strain KD6 isolated from petroleum refinery waste.

Polycyclic aromatic hydrocarbons (PAHs) are a group of environmental pollutant that are given top priority to maintain water and soil quality to the most amenable standard. Biodegradation of PAHs by bacteria is the convenient option for decontamination on site or off site. The aim of the present study was to isolate and identify naturally occurring bacteria having mixed PAHs biodegradation ability. The newly isolated Pseudomonas putida strain KD6 was found to efficiently degrade 97.729% of 1500 mg L-1 mixed PAHs within 12 days in carbon-deficient minimal medium (CSM). The half-life (t1/2) and degradation rate constant (k) were estimated to be 3.2 and 0.2165 days, respectively. The first-order kinetic parameters in soil by strain KD6 had shown efficient biodegradation potency with the higher concentration of total PAHs (1500 mg kg-1 soil), t1/2 = 10.44 days-1. However, the biodegradation by un-inoculated control soil was found slower (t1/2 = 140 days-1) than the soil inoculated with P. putida strain KD6. The enzyme kinetic constants are also in agreement with chemical data obtained from the HPLC analysis. In addition, the sequence analysis and molecular docking studies showed that the strain KD6 encodes a mutant version of naphthalene 1,2-dioxygenase which have better Benzpyrene binding energy (-9.90 kcal mol-1) than wild type (-8.18 kcal mol-1) enzyme (chain A, 1NDO), respectively, with 0.00 and 0.08 RMSD values. The mutated naphthalene 1,2-dioxygenase nahAc has six altered amino acid residues near to the ligand binding site. The strain KD6 could be a good bioresource for in situ or ex situ biodegradation of polycyclic aromatic hydrocarbon.