EPSP synthase

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EPSP Synthase (3-phosphoshikimate 1-carboxyvinyltransferase)
2AA9 PDB.png
Ribbon diagram of EPSP synthase liganded with shikimate (spheres). [1]
Identifiers
EC no. 2.5.1.19
CAS no. 9068-73-9
Databases
IntEnz IntEnz view
BRENDA BRENDA entry
ExPASy NiceZyme view
KEGG KEGG entry
MetaCyc metabolic pathway
PRIAM profile
PDB structures RCSB PDB PDBe PDBsum
Gene Ontology AmiGO / QuickGO
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PMC articles
PubMed articles
NCBI proteins
EPSP synthase (3-phosphoshikimate 1-carboxyvinyltransferase)
Identifiers
SymbolEPSP_synthase
Pfam PF00275
InterPro IPR001986
PROSITE PDOC00097
SCOP2 1eps / SCOPe / SUPFAM
Available protein structures:
Pfam   structures / ECOD  
PDB RCSB PDB; PDBe; PDBj
PDBsum structure summary

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

Contents

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:

Structure

EPSP synthase is a monomeric enzyme with a molecular mass of approximately 46,000. [2] [3] [4] 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. [5]

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. [6] [7] Gut flora of some animals contain EPSPS. [8]

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. [9] [10] 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. [11]

EPSPreactionII.svg

Studies of the enzyme kinetics for this reaction have determined the specific sequence and energetics of each step of the process. [12] 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. [13] 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. [5] [14]

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. [5] [15]

Related Research Articles

<span class="mw-page-title-main">Herbicide</span> Type of chemical used to kill unwanted plants

Herbicides, also commonly known as weed killers, are substances used to control undesired plants, also known as weeds. Selective herbicides control specific weed species while leaving the desired crop relatively unharmed, while non-selective herbicides kill plants indiscriminately. The combined effects of herbicides, nitrogen fertilizer, and improved cultivars has increased yields of major crops by 3x to 6x from 1900 to 2000.

<span class="mw-page-title-main">Glyphosate</span> Systemic herbicide and crop desiccant

Glyphosate is a broad-spectrum systemic herbicide and crop desiccant. It is an organophosphorus compound, specifically a phosphonate, which acts by inhibiting the plant enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSP). It is used to kill weeds, especially annual broadleaf weeds and grasses that compete with crops. Its herbicidal effectiveness was discovered by Monsanto chemist John E. Franz in 1970. Monsanto brought it to market for agricultural use in 1974 under the trade name Roundup. Monsanto's last commercially relevant United States patent expired in 2000.

Shikimic acid, more commonly known as its anionic form shikimate, is a cyclohexene, a cyclitol and a cyclohexanecarboxylic acid. It is an important biochemical metabolite in plants and microorganisms. Its name comes from the Japanese flower shikimi, from which it was first isolated in 1885 by Johan Fredrik Eykman. The elucidation of its structure was made nearly 50 years later.

<span class="mw-page-title-main">Chorismic acid</span> Chemical compound

Chorismic acid, more commonly known as its anionic form chorismate, is an important biochemical intermediate in plants and microorganisms. It is a precursor for:

<span class="mw-page-title-main">Phosphoenolpyruvic acid</span> Chemical compound

Phosphoenolpyruvate is the carboxylic acid derived from the enol of pyruvate and phosphate. It exists as an anion. PEP is an important intermediate in biochemistry. It has the highest-energy phosphate bond found in organisms, and is involved in glycolysis and gluconeogenesis. In plants, it is also involved in the biosynthesis of various aromatic compounds, and in carbon fixation; in bacteria, it is also used as the source of energy for the phosphotransferase system.

<span class="mw-page-title-main">Cyanidin</span> Anthocyanidin pigment in flowering plant petals and fruits

Cyanidin is a natural organic compound. It is a particular type of anthocyanidin. It is a pigment found in many red berries including grapes, bilberry, blackberry, blueberry, cherry, chokeberry, cranberry, elderberry, hawthorn, loganberry, açai berry and raspberry. It can also be found in other fruits such as apples and plums, and in red cabbage and red onion. It has a characteristic reddish-purple color, though this can change with pH; solutions of the compound are red at pH < 3, violet at pH 7-8, and blue at pH > 11. In certain fruits, the highest concentrations of cyanidin are found in the seeds and skin. Cyanidin has been found to be a potent sirtuin 6 (SIRT6) activator.

<span class="mw-page-title-main">Phosphoenolpyruvate carboxylase</span> Class of enzymes

Phosphoenolpyruvate carboxylase (also known as PEP carboxylase, PEPCase, or PEPC; EC 4.1.1.31, PDB ID: 3ZGE) is an enzyme in the family of carboxy-lyases found in plants and some bacteria that catalyzes the addition of bicarbonate (HCO3) to phosphoenolpyruvate (PEP) to form the four-carbon compound oxaloacetate and inorganic phosphate:

<span class="mw-page-title-main">Amino acid synthesis</span> The set of biochemical processes by which amino acids are produced

Amino acid biosynthesis is the set of biochemical processes by which the amino acids are produced. The substrates for these processes are various compounds in the organism's diet or growth media. Not all organisms are able to synthesize all amino acids. For example, humans can synthesize 11 of the 20 standard amino acids. These 11 are called the non-essential amino acids.

<span class="mw-page-title-main">Aromatic amino acid</span> Amino acid having an aromatic ring

An aromatic amino acid is an amino acid that includes an aromatic ring.

<span class="mw-page-title-main">Erythrose 4-phosphate</span> Chemical compound

Erythrose 4-phosphate is a phosphate of the simple sugar erythrose. It is an intermediate in the pentose phosphate pathway and the Calvin cycle.

<span class="mw-page-title-main">Shikimate dehydrogenase</span> Enzyme involved in amino acid biosynthesis

In enzymology, a shikimate dehydrogenase (EC 1.1.1.25) is an enzyme that catalyzes the chemical reaction

<span class="mw-page-title-main">3-dehydroquinate synthase</span> Enzyme

The enzyme 3-dehydroquinate synthase catalyzes the chemical reaction

<span class="mw-page-title-main">Chorismate synthase</span>

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<span class="mw-page-title-main">Shikimate kinase</span> Class of enzymes

Shikimate kinase (EC 2.7.1.71) is an enzyme that catalyzes the ATP-dependent phosphorylation of shikimate to form shikimate 3-phosphate. This reaction is the fifth step of the shikimate pathway, which is used by plants and bacteria to synthesize the common precursor of aromatic amino acids and secondary metabolites. The systematic name of this enzyme class is ATP:shikimate 3-phosphotransferase. Other names in common use include shikimate kinase (phosphorylating), and shikimate kinase II.

The Aminoshikimate pathway is a biochemical pathway present in some plants, which has been studied by biologists, biochemists and especially those interested in manufacture of novel antibiotic drugs. The pathway is a novel variation of the shikimate pathway. The aminoshikimate pathway was first discovered and studied in the rifamycin B producer Amycolatopsis mediterranei. Its end product, 3-amino-5-hydroxybenzoate, serves as an initiator for polyketide synthases in the biosynthesis of ansamycins.

<span class="mw-page-title-main">Shikimate pathway</span> Biosynthetic Pathway

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<span class="mw-page-title-main">DAHP synthase</span> Class of enzymes

3-Deoxy-D-arabinoheptulosonate 7-phosphate (DAHP) synthase is the first enzyme in a series of metabolic reactions known as the shikimate pathway, which is responsible for the biosynthesis of the amino acids phenylalanine, tyrosine, and tryptophan. Since it is the first enzyme in the shikimate pathway, it controls the amount of carbon entering the pathway. Enzyme inhibition is the primary method of regulating the amount of carbon entering the pathway. Forms of this enzyme differ between organisms, but can be considered DAHP synthase based upon the reaction that is catalyzed by this enzyme.

3-Deoxy-<small>D</small>-<i>arabino</i>-heptulosonic acid 7-phosphate Chemical compound

3-Deoxy-D-arabino-heptulosonic acid 7-phosphate (DAHP) is a 7-carbon ulosonic acid. This compound is found in the shikimic acid biosynthesis pathway and is an intermediate in the production of aromatic amino acids.

References

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  2. Goldsbrough P (1990). "Gene amplification in glyphosate-tolerant tobacco cells". Plant Science. 72 (1): 53–62. doi:10.1016/0168-9452(90)90186-r.
  3. 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.
  4. 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.
  5. 1 2 3 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.
  6. 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.
  7. 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.
  8. 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. doi:10.2134/jeq2005.0378. PMID   16899736.
  9. 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.
  10. 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.
  11. 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.
  12. 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.
  13. 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. doi: 10.1039/D0RA09061A . PMC   9058485 . PMID   35517162.
  14. 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.
  15. 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. doi:10.1021/jf101286h. PMC   3105486 . PMID   20586458.

Further reading