EPSP synthase - Misplaced Pages

5-enolpyruvylshikimate-3-phosphate (EPSP) synthase is an enzyme produced by plants and microorganisms. EPSPS catalyzes the chemical reaction:

phosphoenolpyruvate (PEP) + 3-phospho shikimate (S3P) ⇌ phosphate + 5-enolpyruvylshikimate-3-phosphate (EPSP)

Thus, the two substrates of this enzyme are phosphoenolpyruvate (PEP) and 3-phosphoshikimate, whereas its two products are phosphate and 5-enolpyruvylshikimate-3-phosphate.

This enzyme is absent from animal genomes, making it an attractive target for herbicides such as glyphosate. A glyphosate-resistant version of the enzyme's gene has been incorporated into genetically modified crops.

Nomenclature

The enzyme belongs to the family of transferases, to be specific those transferring aryl or alkyl groups other than methyl groups. The systematic name of this enzyme class is phosphoenolpyruvate:3-phosphoshikimate 5-O-(1-carboxyvinyl)-transferase. Other names in common use include:

5-enolpyruvylshikimate-3-phosphate synthase,
3-enolpyruvylshikimate 5-phosphate synthase,
3-enolpyruvylshikimic acid-5-phosphate synthetase,
5′-enolpyruvylshikimate-3-phosphate synthase,
5-enolpyruvyl-3-phosphoshikimate synthase,
5-enolpyruvylshikimate-3-phosphate synthetase,
5-enolpyruvylshikimate-3-phosphoric acid synthase,
enolpyruvylshikimate phosphate synthase, and
3-phosphoshikimate 1-carboxyvinyl transferase.

Structure

EPSP synthase is a monomeric enzyme with a molecular mass of approximately 46,000. It consists of two domains connected by protein strands that function as a hinge, allowing the two domains to move closer together. When a substrate binds to the enzyme, the conformational change causes the domains to clamp around the substrate at the active site.

EPSP synthase is classified into two groups based on sensitivity to glyphosate. Class I enzymes, found in plants and some bacteria, are inhibited by low micromolar concentrations of glyphosate. Class II enzymes, found in other bacteria, are resistant to glyphosate inhibition.

Shikimate pathway

EPSP synthase participates in the biosynthesis of the aromatic amino acids phenylalanine, tyrosine, and tryptophan via the shikimate pathway in bacteria, fungi, and plants. EPSP synthase is produced only by plants and micro-organisms; the gene coding for it is not in the mammalian genome. Gut flora of some animals contain EPSPS.

Reaction

EPSP synthase catalyzes the reaction which converts shikimate-3-phosphate plus phosphoenolpyruvate to 5-enolpyruvylshikimate-3-phosphate (EPSP) by way of an acetal-like tetrahedral intermediate. Basic and acidic amino acids in the active site are involved in deprotonation of the hydroxyl group of PEP and in the proton-exchange steps related to the tetrahedral intermediate itself, respectively.

Studies of the enzyme kinetics for this reaction have determined the specific sequence and energetics of each step of the process. A neutrally charged lysine (lys-22) acts as a general base, deprotonating the hydroxyl group of S3P such that the resulting oxyanion can attack the most electrophilic carbon atom of PEP. A glutamic acid (glu-341) acts as a general acid by donating a proton. The deprotonated glu-341 then acts as a base, taking back its proton, and the S3P group is kicked off and protonated by the protonated lysine.

Herbicide target

EPSP synthase is the biological target for the herbicide glyphosate. Glyphosate is a competitive inhibitor of EPSP synthase, acting as a transition state analog that binds more tightly to the EPSPS-S3P complex than PEP and inhibits the shikimate pathway. This binding leads to inhibition of the enzyme's catalysis and shuts down the pathway. Eventually this results in organism death from lack of aromatic amino acids the organism requires to survive.

A version of the enzyme that both was resistant to glyphosate and that was still efficient enough to drive adequate plant growth was identified by Monsanto scientists after much trial and error in an Agrobacterium strain called CP4 (Q9R4E4). The strain CP4 was found surviving in a waste-fed column at a glyphosate production facility. The CP4 EPSP synthase enzyme has been engineered into several genetically modified crops.

References

Priestman MA, Healy ML, Funke T, Becker A, Schönbrunn E (October 2005). "Molecular basis for the glyphosate-insensitivity of the reaction of 5-enolpyruvylshikimate 3-phosphate synthase with shikimate". FEBS Letters. 579 (25): 5773–5780. Bibcode:2005FEBSL.579.5773P. doi:10.1016/j.febslet.2005.09.066. PMID 16225867. S2CID 26614581.
Goldsbrough P (1990). "Gene amplification in glyphosate-tolerant tobacco cells". Plant Science. 72 (1): 53–62. Bibcode:1990PlnSc..72...53G. doi:10.1016/0168-9452(90)90186-r.
Abdel-Meguid SS, Smith WW, Bild GS (December 1985). "Crystallization of 5-enolpyruvylshikimate 3-phosphate synthase from Escherichia coli". Journal of Molecular Biology. 186 (3): 673. doi:10.1016/0022-2836(85)90140-8. PMID 3912512.
Ream JE, Steinrücken HC, Porter CA, Sikorski JA (May 1988). "Purification and Properties of 5-Enolpyruvylshikimate-3-Phosphate Synthase from Dark-Grown Seedlings of Sorghum bicolor". Plant Physiology. 87 (1): 232–238. doi:10.1104/pp.87.1.232. PMC 1054731. PMID 16666109.
^ Pollegioni L, Schonbrunn E, Siehl D (August 2011). "Molecular basis of glyphosate resistance-different approaches through protein". The FEBS Journal. 278 (16): 2753–2766. doi:10.1111/j.1742-4658.2011.08214.x. PMC 3145815. PMID 21668647.
Funke T, Han H, Healy-Fried ML, Fischer M, Schönbrunn E (August 2006). "Molecular basis for the herbicide resistance of Roundup Ready crops". Proceedings of the National Academy of Sciences of the United States of America. 103 (35): 13010–13015. Bibcode:2006PNAS..10313010F. doi:10.1073/pnas.0603638103. JSTOR 30050705. PMC 1559744. PMID 16916934.
Maeda H, Dudareva N (2012). "The shikimate pathway and aromatic amino Acid biosynthesis in plants". Annual Review of Plant Biology. 63 (1): 73–105. doi:10.1146/annurev-arplant-042811-105439. PMID 22554242. The AAA pathways consist of the shikimate pathway (the prechorismate pathway) and individual postchorismate pathways leading to Trp, Phe, and Tyr.... These pathways are found in bacteria, fungi, plants, and some protists but are absent in animals. Therefore, AAAs and some of their derivatives (vitamins) are essential nutrients in the human diet, although in animals Tyr can be synthesized from Phe by Phe hydroxylase....The absence of the AAA pathways in animals also makes these pathways attractive targets for antimicrobial agents and herbicides.
Cerdeira AL, Duke SO (2006). "The current status and environmental impacts of glyphosate-resistant crops: a review". Journal of Environmental Quality. 35 (5): 1633–1658. Bibcode:2006JEnvQ..35.1633C. doi:10.2134/jeq2005.0378. PMID 16899736.
Furdui CM, Anderson KS (2010). "8.18.4.1.1. EPSP synthase: A tetrahedral ketal phosphate enzyme intermediate". Comprehensive Natural Products II. Chemistry and Biology. Reference Module in Chemistry, Molecular Sciences and Chemical Engineering. Vol. 8. pp. 663–688. doi:10.1016/B978-008045382-8.00158-1.
Anderson KS, Sammons RD, Leo GC, Sikorski JA, Benesi AJ, Johnson KA (February 1990). "Observation by 13C NMR of the EPSP synthase tetrahedral intermediate bound to the enzyme active site". Biochemistry. 29 (6): 1460–1465. doi:10.1021/bi00458a017. PMID 2334707.
Park H, Hilsenbeck JL, Kim HJ, Shuttleworth WA, Park YH, Evans JN, et al. (February 2004). "Structural studies of Streptococcus pneumoniae EPSP synthase in unliganded state, tetrahedral intermediate-bound state and S3P-GLP-bound state". Molecular Microbiology. 51 (4): 963–971. doi:10.1046/j.1365-2958.2003.03885.x. PMID 14763973. S2CID 45549442.
Anderson KS, Sikorski JA, Johnson KA (September 1988). "A tetrahedral intermediate in the EPSP synthase reaction observed by rapid quench kinetics". Biochemistry. 27 (19): 7395–7406. doi:10.1021/bi00419a034. PMID 3061457.
Fonseca EC, da Costa KS, Lameira J, Alves CN, Lima AH (December 2020). "Investigation of the target-site resistance of EPSP synthase mutants P106T and T102I/P106S against glyphosate". RSC Advances. 10 (72): 44352–44360. Bibcode:2020RSCAd..1044352F. doi:10.1039/D0RA09061A. PMC 9058485. PMID 35517162.
Schönbrunn E, Eschenburg S, Shuttleworth WA, Schloss JV, Amrhein N, Evans JN, et al. (February 2001). "Interaction of the herbicide glyphosate with its target enzyme 5-enolpyruvylshikimate 3-phosphate synthase in atomic detail". Proceedings of the National Academy of Sciences of the United States of America. 98 (4): 1376–1380. Bibcode:2001PNAS...98.1376S. doi:10.1073/pnas.98.4.1376. PMC 29264. PMID 11171958.
Green JM, Owen MD (June 2011). "Herbicide-resistant crops: utilities and limitations for herbicide-resistant weed management". Journal of Agricultural and Food Chemistry. 59 (11): 5819–5829. Bibcode:2011JAFC...59.5819G. doi:10.1021/jf101286h. PMC 3105486. PMID 20586458.

v t e Enzymes
Activity	Active site Binding site Catalytic triad Oxyanion hole Enzyme promiscuity Diffusion-limited enzyme Cofactor Enzyme catalysis
Regulation	Allosteric regulation Cooperativity Enzyme inhibitor Enzyme activator
Classification	EC number Enzyme superfamily Enzyme family List of enzymes
Kinetics	Enzyme kinetics Eadie–Hofstee diagram Hanes–Woolf plot Lineweaver–Burk plot Michaelis–Menten kinetics
Types	EC1 Oxidoreductases (list) EC2 Transferases (list) EC3 Hydrolases (list) EC4 Lyases (list) EC5 Isomerases (list) EC6 Ligases (list) EC7 Translocases (list)