Abstract
Study of enzymes in halophiles will help to understand the mechanism of aromatic hydrocarbons degradation in saline environment. In this study, two novel catechol 2,3-dioxygenases (C23O1 and C23O2) were cloned and overexpressed from a halophilic bacterial consortium enriched from an oil-contaminated saline soil. Phylogenetic analysis indicated that the novel C23Os and their relatives formed a new branch in subfamily I.2.A of extradiol dioxygenases and the sequence differences were further analyzed by amino acid sequence alignment. Two enzymes with the halotolerant feature were active over a range of 0–30% salinity and they performed more stable at high salinity than in the absence of salt. Surface electrostatic potential and amino acids composition calculation suggested high acidic residues content, accounting for their tolerance to high salinity. Moreover, two enzymes were further characterized. The enzymes activity both increased in the presence of Fe3+, Fe2+, Cu2+ and Al3+ and showed no significant inhibition by other tested metal ions. The optimal temperatures for the C23Os were 40 °C and 60 °C and their best substrates were catechol and 4-methylcatechol respectively. As the firstly isolated and characterized catechol dioxygenases from halophiles, the two halotolerant C23Os presented novel characteristics suggesting their potential application in aromatic hydrocarbons biodegradation.
Highlights
Concerning the genes and enzymes involved in aromatic hydrocarbons degradation in halophiles
As a crucial enzyme involved in the degradation of aromatic hydrocarbons, catechol 2, 3-dioxygenase (C23O, key enzyme of meta-cleavage pathway) has been identified as one of the most widespread ring cleavage enzymes[22] and many C23Os have been reported from non-halophilic bacteria (Pseudomonas, Sphingomonas, Acinetobacter, Bacillus, Ralstonia, Burkholderia and Nocardia)[23]
A dynamic pattern of C23O activity of this bacterial consortium was measured during degradation of phenanthrene at 10% salinity (Fig. S1)
Summary
Concerning the genes and enzymes involved in aromatic hydrocarbons degradation in halophiles. Kim et al.[20] and Moreno et al.[21] futher separately studied the genetic organization of benzoate and benzene-degrading gene clusters from two halophiles to elucidate the compounds degradation pathway. As a crucial enzyme involved in the degradation of aromatic hydrocarbons, catechol 2, 3-dioxygenase (C23O, key enzyme of meta-cleavage pathway) has been identified as one of the most widespread ring cleavage enzymes[22] and many C23Os have been reported from non-halophilic bacteria (Pseudomonas, Sphingomonas, Acinetobacter, Bacillus, Ralstonia, Burkholderia and Nocardia)[23]. This study helps to develop the knowledge of the role of halotolerant enzymes in aromatic hydrocarbons degradation by halophilic bacteria and to understand the specific properties presented by halotolerant catechol dioxygenases for potential application
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