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Topramezone is a 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitor. Due to its broad-spectrum, high efficiency, and low toxicity, topramezone is a candidate herbicide for the construction of genetically modified (GM) herbicide-resistant crops. In the present study, we screened a topramezone-resistant isolate Sphingobium sp. TPM-19 and cloned a topramezone-resistant HPPD gene ( SphppD ) from this isolate. Sp HPPD shared the highest similarity (53%) with an HPPD from Vibrio vulnificus CMCP6. Sp HPPD was synthesized in Escherichia coli BL21(DE3) and purified to homogeneity using Co 2+ -affinity chromatography. Sp HPPD was found to be a monomer. The K m and k cat of Sp HPPD for 4-hydroxyphenylpyruvate (4-HPP) were 82.8 μM and 15.0 s -1 , respectively. Sp HPPD showed high resistance to topramezone with half maximal inhibitory concentration (IC 50 ) and K i values of 5.2 and 2.5 μM, respectively. Additionally, Sp HPPD also showed high resistance to isoxaflutole (DKN) (IC 50 : 8.7 μM; K i : 6.0 μM) and mesotrione (IC 50 : 4.2 μM; K i : 1.3 μM) and moderate resistance to tembotrione (IC 50 : 2.5 μM; K i : 1.0 μM). The introduction of the SphppD gene into Arabidopsis thaliana enhanced obvious resistance against topramezone. In conclusion, this study provides a novel topramezone-resistant HPPD gene for the genetic engineering of GM herbicide-resistant crops.

Esterase SulE detoxicates a variety of sulfonylurea herbicides through de-esterification. SulE exhibits high activity against thifensulfuron-methyl but low activity against other sulfonylureas. In this study, two variants, m2311 (P80R) and m0569 (P80R and G176A), with improved activity were screened from a mutation library constructed by error-prone PCR. Variant m2311 showed a higher activity against sulfonylureas in comparison variant m0569 and was further investigated. The k cat / K m value of variant m2311 for metsulfuron-methyl, sulfometuron-methyl, chlorimuron-ethyl, tribenuron-methyl, and ethametsulfuron-methyl increased by 3.20-, 1.72-, 2.94-, 2.26- and 2.96-fold, respectively, in comparison with the wild type. Molecular modeling suggested that the activity improvement of variant m2311 is due to the substitution of Pro80 by arginine, leading to the formation of new hydrogen bonds between the enzyme and substrate. This study facilitates further elucidation of the structure and function of SulE and provides an improved gene resource for the detoxification of sulfonylurea residues and the genetic engineering of sulfonylurea-resistant crops.