Enhancement of haloacetate dehalogenase production by strain mutation and condition optimization

Abstract

Enhancement of the activity of an inducible chloroacetate dehalogenase was carried out by efficient and safe mutation with UV and microwave irradiation along with optimization of culture conditions. First, a stable mutant of Pseudomonas sp. CGMCC 3267-MW6 with chloroacetate dehalogenase activity of 2.77 U/mL (3-fold higher activity than the wild strain) was produced by mutation. The maximum activity of this inducible enzyme was measured as 29.41 U/mL when Pseudomonas sp. CGMCC 3267-MW6 was cultured with 4 g/L 3-hydroxybutyrate for 12 h followed by 40 mM 3-chlorobutyrate for an additional 20 h. Production of the enzyme was found to be associated with growth of the bacterium. According to these results, we determined that the optimum inducer of chloroacetate dehalogenase activity would be a hard degradable substrate. The optimum auxiliary carbon source would be the primary metabolite of the substrate or the precursor of the metabolite. The optimum time of inducer supplementation would be during the middle stage of exponential phase. The optimum concentration of substrate would be sufficient but would not induce inhibition. Finally, the optimum collection time would be at the later stage of exponential phase. This work provides further knowledge of chloroacetate dehalogenase and the optimization of inducible enzyme production.

Implications of microbial adaptation for the assessment of environmental persistence of chemicals

“…Even if beneficial mutations are rare and can be lost by successive replication and random drift (Elena & Lenski, 2003), they can accumulate into the genetic material of the population over time. New catabolic pathways can be produced and production of specific enzymes can be increased by the accumulation of mutations (Lin, Yang, Xu, & Wu, 2011;Madigan et al, 2014). The implication of mutations into the dynamics of evolutionary adaption has been well documented (Elena & Lenski, 2003).…”

Ochrobactrum teleogrylli sp. nov., a pesticide-degrading bacterium isolated from the insect Teleogryllus occipitalis living in deserted cropland

“…They have been widely used for the biodegradation of haloacetates, especially chloroacetate. Additionally, they can potentially be used for the preparation of 2-hydroxyalkanic acid [34]. However, some gene families that are shared by Ochrobactrum pecoris 08RB2639 T and Ochrobactrum haematophilum CCUG 38531 T were lacking in LCB8 T .…”

Enhancement of L-2-haloacid dehalogenase expression in Pseudomonas stutzeri DEH138 based on the different substrate specificity between dehalogenase-producing bacteria and their dehalogenases

“…Besides DEH138, the inconsistency between halide degradation activity with different bacteria and their corresponding dehalogenases were also observed with Pseudomonas putida P3 and S3 (Slater et al 1979), Pseudomonas sp. CGMCC 3267-MW6 (Lin et al 2011), X. autotrophicus GJ10 (Janssen et al 1985) and Rhizobium sp. RC1 and their corresponding enzymes (Cairns et al 1996).…”