Molecular basis for the glyphosate‐insensitivity of the reaction of 5‐enolpyruvylshikimate 3‐phosphate synthase with shikimate

Priestman, Melanie A.; Healy, Martha L.; Funke, T.; Becker, Andreas; Schönbrunn, E.

doi:10.1016/j.febslet.2005.09.066

Cited by 23 publications

(13 citation statements)

References 21 publications

(28 reference statements)

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“…Indeed, EPSPS activity was found in an in vitro assay in P. aegyptiaca flowering shoots and callus extracts for both the enzyme substrate S3P and its precursor shikimate. Although the natural substrate of EPSPS is S3P, the enzyme may also utilize shikimate as a substrate; however, this last reaction is less sensitive to glyphosate (Priestman et al, 2005) and indeed, in our experiments, ID 50 for EPSPS extracted from the callus with S3P as substrate was 100 times lower than the same reaction using shikimate as the substrate ( Figure 7 ).…”

Section: Discussionmentioning

confidence: 62%

“…EPSPS extracted from flowering shoots was much more sensitive to the herbicide than the enzyme extracted from callus, with respective ID 50 values of about 84 ± 0.03 and 794 ± 35 μM, using shikimate as the substrate ( Figure 7A ). The enzymatic reaction with this substrate is less sensitive to glyphosate (Priestman et al, 2005) than the same reaction with the natural substrate S3P. With the latter, the ID 50 value for EPSPS from callus was about 8 ± 0.29 μM ( Figure 7B ).…”

Section: Resultsmentioning

confidence: 93%

“…5-enolpyruvylshikimate-3-phosphate synthase extraction and assay were based on the methods of Eschenburg et al (2002) and Priestman et al (2005) with modifications. Briefly, callus or tips of young flowering shoots were extracted in two volumes (w/v) of extraction buffer (50 mM Tris–HCl pH 7.8, 1 mM EDTA, 0.1 M NaCl, 1 mM DTT, 0.1% v/v Triton X-100) with lysozyme (1 mg lysozyme g -1 plant tissue).…”

Section: Methodsmentioning

confidence: 99%

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The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape

et al. 2017

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It is not clear why herbicides targeting aromatic and branched-chain amino acid biosynthesis successfully control broomrapes—obligate parasitic plants that obtain all of their nutritional requirements, including amino acids, from the host. Our objective was to reveal the mode of action of imazapic and glyphosate in controlling the broomrape Phelipanche aegyptiaca and clarify if this obligatory parasite has its own machinery for the amino acids biosynthesis. P. aegyptiaca callus was studied to exclude the indirect influence of the herbicides on the parasite through the host plant. Using HRT – tomato plants resistant to imidazolinone herbicides, it was shown that imazapic is translocated from the foliage of treated plants to broomrape attachments on its roots and controls the parasite. Both herbicides inhibited P. aegyptiaca callus growth and altered the free amino acid content. Blasting of Arabidopsis thaliana 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) and acetolactate synthase (ALS) cDNA against the genomic DNA of P. aegyptiaca yielded a single copy of each homolog in the latter, with about 78 and 75% similarity, respectively, to A. thaliana counterparts at the protein level. We also show for the first time that both EPSPS and ALS are active in P. aegyptiaca callus and flowering shoots and are inhibited by glyphosate and imazapic, respectively. Thus leading to deficiency of those amino acids in the parasite tissues and ultimately, death of the parasite, indicating the ability of P. aegyptiaca to synthesize branched-chain and aromatic amino acids through the activity of ALS and EPSPS, respectively.

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Section: Discussionmentioning

confidence: 62%

Section: Resultsmentioning

confidence: 93%

Section: Methodsmentioning

confidence: 99%

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The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape

et al. 2017

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“…Development of glyphosate-tolerant crops was a major breakthrough in agricultural biotechnology. Recent advances have been made in identifying, cloning, and testing the glyphosateinsensitive form of EPSPS enzymes [3,8,22,23]. In these reports, EPSP synthases have been divided into two classes, sharing less than 30% amino acid identity.…”

Section: Introductionmentioning

confidence: 99%

Functional Characterization of aroA from Rhizobium leguminosarum with Significant Glyphosate Tolerance in Transgenic Arabidopsis

Han

Tian²,

Xu³

et al. 2014

Journal of Microbiology and Biotechnology

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Glyphosate is the active component of the top-selling herbicide, the phytotoxicity of which is due to its inhibition of the shikimic acid pathway. 5-Enolpyruvylshikimate-3-phosphate synthase (EPSPS) is a key enzyme in the shikimic acid pathway. Glyphosate tolerance in plants can be achieved by the expression of a glyphosate-insensitive aroA gene (EPSPS). In this study, we used a PCR-based two-step DNA synthesis method to synthesize a new aroA gene (aroA R. leguminosarum) from Rhizobium leguminosarum. In vitro glyphosate sensitivity assays showed that aroA R. leguminosarum is glyphosate tolerant. The new gene was then expressed in E. coli and key kinetic values of the purified enzyme were determined. Furthermore, we transformed the aroA gene into Arabidopsis thaliana by the floral dip method. Transgenic Arabidopsis with the aroA R. leguminosarum gene was obtained to prove its potential use in developing glyphosate-resistant crops.

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“…Numerosos estudos usando diferentes técnicas demonstraram que o glifosato forma preferencialmente um complexo ternário estável com a enzima EPSPs e S3P (EPSP-S3P-glifosato), este complexo ternário representa a forma atualmente aceita como responsável pela atividade do herbicida (Feng et al, 2005;Herrmann & Weaver, 1999;Priestman, Healy, Funke, Becker, & Schönbrunn, 2005).…”

Section: Modo E Mecanismo De Açãounclassified

Modelagem molecular de quitosana e glifosato e análise de suas interações

Faria¹

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pela irmandade e exemplos. Ao Prof. Dr. Moacyr Comar pela participação na banca e por contribuir na qualificação. Ao Prof. Dr. Guedmiller S. Oliveira pela participação na banca e por contribuir na qualificação. Ao Prof. Dr. Osmando Ferreira por contribuir na qualificação. Ao Prof. Dr. Erick França por todas as gentilezas e companheirismo. À Profa. Dra. Gislaine Fernandes pela chefia e exemplo durante a carreira profissional. Ao Prof. Dr. Paulo Henrique Soares por me oferecer além de abrigo, ensinamentos para toda a vida. Ao Prof. Dr. Rodrigo Muñoz pelo exemplo profissional e motivacional. Ao Dr. Moacir Junior pelos seus grandes exemplos durante seu doutorado. Ao Me. Lourival Rodrigues de Sousa Neto por acompanhar toda minha caminhada. Ao Me. Renan Faria Guerra pela parceria e exemplos. Ao Prof. Me. Bruno Pereira Garcês por me acompanhar desde 2007 no início da graduação. Ao meu pai pelo pilar e todos os princípios ensinados. À minha mãe por ser amiga, companheira e incrível. Às minhas irmãs pela força e carinho. À minha namorada pela paciência e companheirismo. À minha família que é minha fortaleza. À minha avó pela proteção eterna. Aos meus grandes amigos Guilherme Freitas,

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Molecular basis for the glyphosate‐insensitivity of the reaction of 5‐enolpyruvylshikimate 3‐phosphate synthase with shikimate

Cited by 23 publications

References 21 publications

The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape

The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape

Functional Characterization of aroA from Rhizobium leguminosarum with Significant Glyphosate Tolerance in Transgenic Arabidopsis

Modelagem molecular de quitosana e glifosato e análise de suas interações

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