A T42M Substitution in Bacterial 5-Enolpyruvylshikimate-3-phosphate Synthase (EPSPS) Generates Enzymes with Increased Resistance to Glyphosate

He, Meiqi; Nie, Yan-Fang; Xu, Peng

doi:10.1271/bbb.67.1405

Cited by 34 publications

(24 citation statements)

References 14 publications

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“…Resistant biotypes of E. indica carrying two mutations in their EPSPS were recently reported , Yu et al 2015. The ability to withstand glyphosate has also been conferred to plants by introducing point mutations in the EPSPS gene through site-directed mutagenesis, gene targeting, and DNA shuffling (He et al 2003;Tian et al 2013;Zhang et al 2011). Naturally glyphosatetolerant Allium macrostemon has an EPSPS that transformed and expressed into E. coli, and tobacco confers them resistance to the herbicide Zhou et al 2009).…”

Section: Discussionmentioning

confidence: 99%

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

Mao

Xie

Chen

et al. 2015

Planta

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A combination of unique EPSPS structure and increased gene copy number and expression contribute to natural glyphosate tolerance in three lilyturf species. A few plants are naturally tolerant to glyphosate, the most widely used non-selective herbicide worldwide. Here, the basis for natural tolerance to glyphosate in three lilyturf species, Ophiopogon japonicus (OJ), Liriope spicata (LS), and Liriope platyphylla (LP), is characterized. These species tolerate glyphosate at about five times the commercially recommended field dose. They share three unique amino acids in their 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) that affect glyphosate binding. These correspond to Asp71Met, Ala112Ile, and Val201Met amino acid variations compared to 231 other published plant EPSPS amino acid sequences. There was also a common deletion at 91 of a highly conserved glutamic acid. Glyphosate-treated lilyturf plants accumulated little shikimic acid but had significantly higher levels of EPSPS mRNA than initially expressed in the control. The IC50 of LsEPSPS was 14.0 µM compared to the 5.1 µM of Arabidopsis thaliana. The higher K m and K i values of LsEPSPS kinetics showed that LsEPSPS had lower substrate binding affinity to glyphosate. Overexpression of LsEPSPS in the recombinant E. coli BL21 (DE3) strain enhanced its tolerance to glyphosate. Both OJ and LS had two copies of the EPSPS gene, while LP had three copies. Therefore, a combination of unique EPSPS structure and increased gene copy number and expression contribute to natural glyphosate tolerance in the three lilyturf species.

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Section: Discussionmentioning

confidence: 99%

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

Mao

Xie

Chen

et al. 2015

Planta

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“…Numerous amino acid residues that affect EPSPS glyphosate resistance have already been identified (1,8,10,13,15,19,20,24,25,31,35,40,50). Notably, these residues are mostly distributed in two regions according to their 3D structure.…”

Section: Discussionmentioning

confidence: 99%

“…One region is at the hinge between the two large globular domains of EPSPS. A typical example in this region is the T42M substitution of bacterial EPSPS that generates an enzyme with an increased resistance to glyphosate (13). The other is the third helix of the core of the N-terminal domain.…”

Section: Discussionmentioning

confidence: 99%

“…Three new residues-N267S, P318R, and M425T-that also affect glyphosate resistance have been identified in our study. Among them, N267S is located near the hinge between the two EPSPS globular domains (13). However, the P318R and M425T residues are not located in these two regions, but at the upper surface of the lower hemisphere in the interdomain cleft of the closed enzyme state.…”

Section: Discussionmentioning

confidence: 99%

“…In the case of directed evolution targeting the glyphosate resistance of EPSPS, both He et al and Zhou et al also obtained mutant EPSPS with increased glyphosate resistance and enzyme activity (13,50). Therefore, mutant EPSPSs with improved enzymatic parameters such as higher K i (glyphosate) and normal K m (PEP) values can be obtained by DNA shuffling.…”

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Improvement of Glyphosate Resistance through Concurrent Mutations in Three Amino Acids of the Ochrobactrum 5-Enopyruvylshikimate-3-Phosphate Synthase

Tian

Xiong

et al. 2011

Appl Environ Microbiol

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A mutant of 5-enopyruvylshikimate-3-phosphate synthase from Ochrobactrum anthropi was identified after four rounds of DNA shuffling and screening. Its ability to restore the growth of the mutant ER2799 cell on an M9 minimal medium containing 300 mM glyphosate led to its identification. The mutant had mutations in seven amino acids: E145G, N163H, N267S, P318R, M377V, M425T, and P438L. Among these mutations, N267S, P318R, and M425T have never been previously reported as important residues for glyphosate resistance. However, in the present study they were found by site-directed mutagenesis to collectively contribute to the improvement of glyphosate tolerance. Kinetic analyses of these three mutants demonstrated that the effectiveness of these three individual amino acid alterations on glyphosate tolerance was in the order P318R > M425T > N267S. The results of the kinetic analyses combined with a three-dimensional structure modeling of the location of P318R and M425T demonstrate that the lower hemisphere's upper surface is possibly another important region for glyphosate resistance. Furthermore, the transgenic Arabidopsis was obtained to confirm the potential of the mutant in developing glyphosate-resistant crops.

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Identification and analysis of 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS) gene from glyphosate‐resistant Ochrobactrum intermedium Sq20

Firdous

Iqbal

Anwar

et al. 2017

Pest Management Science

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A glyphosate-resistant bacterial strain O. intermedium Sq20 was discovered. Sequence analysis and structure modelling demonstrated that AroA closely resembles class II EPSPS and possesses high glyphosate resistance. This provides a good foundation for functional analysis of experimentally derived crystal structures. The cloning and characterization of AroA will further help in understanding its role at the molecular level and its potential use in the production of glyphosate-resistant transgenic crops. © 2017 Society of Chemical Industry.

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A T42M Substitution in Bacterial 5-Enolpyruvylshikimate-3-phosphate Synthase (EPSPS) Generates Enzymes with Increased Resistance to Glyphosate

Cited by 34 publications

References 14 publications

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

Improvement of Glyphosate Resistance through Concurrent Mutations in Three Amino Acids of the Ochrobactrum 5-Enopyruvylshikimate-3-Phosphate Synthase

Identification and analysis of 5‐enolpyruvylshikimate‐3‐phosphate synthase (EPSPS) gene from glyphosate‐resistant Ochrobactrum intermedium Sq20

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