1.14.13.20	F424A	site-directed mutagenesis, analysis of substrate binding by the mutant and docking study in comparison to the wild-type enzyme	765761	
1.14.13.20	H47A	site-directed mutagenesis, analysis of substrate binding by the mutant and docking study in comparison to the wild-type enzyme	765761	
1.14.13.20	I48A	site-directed mutagenesis, analysis of substrate binding by the mutant and docking study in comparison to the wild-type enzyme	765761	
1.14.13.20	additional information	construction of tfdBI and tfdBII inactivation mutants on pJP4 plasmid, constructed in Escherichia coli strain BW25113 by allelic replacement of each of the genes tfdBI or tfdBII with a kanamycin resistance cassette, tfdBI and tfdBII are required for growth on 2,4-dichlorophenoxyacetate and 4-chloro-2-methylphenoxyacetate, with tfdBI contributing to a higher extent than tfdBII, phenotype, overview, accumulation of chlorophenol inhibits chlorophenoxyacetate degradation, and inactivation of the tfdB genes enhances the toxic effect of 2,4-dichlorophenoxyacetate on Cupriavidus necator cells	671456	
1.14.13.20	additional information	enzyme TfdB-JLU is engineered by rational design to further broaden its substrate scope towards chlorophenols (CPs). Dissection of the architectures of enzymes from oxidoreductase families to discover their underlying structural sources of substrate promiscuity, and homology modeling of TfdB-JLU. Docking experiments of this homology model with its natural substrate 2,4-dichlorophenol reveals that the phenyl rings of 2,4-DCP form strong interactions with residues His47, Ile48, Trp222, Pro316, and Phe424. These residues are found to be important for substrate binding in the active site. Site-directed mutagenesis strategy is applied for redesigning substrate promiscuity in enzyme TfdB-JLU. P316 is the key residue for enzyme functional engineering	765761	
1.14.13.20	additional information	overexpression of tdfA inhibits chlorophenoxyacetate degradation in Burkholderia xenovorans/pJP4 cells	671456	
1.14.13.20	additional information	synthesis of hybrid nanoflowers (hNFs) formed from cold-adapted 2,4-dichlorophenol hydroxylase (tfdBJLU) and Cu3(PO4)2 x 3H2O. Analysis of the influence of experimental factors, such as the pH of the solution mixture and the enzyme and Cu2+ concentrations, on the activity of the prepared tfdB-JLU-hNFs. 200 mM is the optimal Cu2+ concentration to get tfdB-JLU-hNFs with highest activity recovery, method optimization, overview. The tfdB-JLU-hNFs exhibit excellent durability with 58.34% residual activity after six successive cycles, and up to 90.58% residual activity after 20 days of storage. Comparison of storage stability of tfdB-JLU-hNFs and free tfdB-JLU at 1 mg/ml. This multistage and hierarchical flower-like structure can effectively increase enzyme activity and stability with respect to those of the free enzyme. The satisfactory removal rate of 2,4-dichlorophenol catalyzed by tfdB-JLU-hNFs suggests that this immobilized enzyme exhibits great potential for application in bioremediation	765760	
1.14.13.20	P316A	site-directed mutagenesis, analysis of substrate binding by the mutant and docking study in comparison to the wild-type enzyme	765761	
1.14.13.20	P316Q	site-directed mutagenesis, the TfdB-JLU variant shows a significant enhancement of activity (up to 3.4fold) toward 10 CP congeners compared to wild-type TfdB-JLU. The active improvements of TfdB-JLU-P316Q toward CP congeners show significant difference, especially for active improvements of positional congeners such as 3-CP (1.1fold) compared to 4-CP (3.0fold), as well as 2,3-DCP (1.2fold) compared to 2,5-DCP (3.4fold). Structural analysis results indicate that the improvement in substrate promiscuity of the variant enzyme compared to the wild-type enzyme is possibly due to the increase of non-bonding interaction	765761	
1.14.13.20	W222A	site-directed mutagenesis, analysis of substrate binding by the mutant and docking study in comparison to the wild-type enzyme	765761