Cofactors

Mg(2+).

Reaction Mechanism

phosphopyruvate hydratase By Reference

Yeast enolase (2-phospho-D-glycerate hydrolase) is a metalloenzyme which catalyses the reversible dehydration of D-2-phos-phoglycerate (PGA) to phosphoenolpyruvate (PEP). The enzyme has an absolute requirement for the presence of a divalent cation, as is characteristic of the enolase family.

Mitochondrial targeting of tRK1 in yeast is achieved by the successive actions of enolase 2 and the precursor of the mitochondrial lysyl-tRNA synthetase (preMSK). At the mitochondrial outer membrane, preMSK takes over enolase to start the import process properly; A fraction of the canonical tRNA L-form tRK1 pool is deviated from the cytosolic translation process by the enolase 2, which favours the tRNA conformational change leading to the formation of the F-form.




• Summary
• Step 1
• Step 2
• Products

This enzyme catalyses the inter-conversion of 2-PGA and PEP in a reversible manner. In the dehydration direction (shown here) both Lys345 and Glu211 (the catalytic acid/base pair) are neutral in charge [PMID:8634301, PMID:12846578]. At the end of the dehydration reaction, the residues are in the correct protonation state to perform the hydration reaction, i.e. Lys345 is positively charged and Glu211 negatively charged. It is assumed that at physiological pH that both protonation states of the enzyme coexist in reasonable proportions [PMID:12846578].

Catalytic Residues

AA	Uniprot	Uniprot Resid	PDB	PDB Resid
Glu	P00924	212	7enl	211
His	P00924	374	7enl	373
Glu	P00924	169	7enl	168
Lys	P00924	397	7enl	396
Lys	P00924	346	7enl	345
Ser	P00924	40	7enl	39
Asp	P00924	247	7enl	246
Glu	P00924	296	7enl	295
Asp	P00924	321	7enl	320
His	P00924	160	7enl	159

Step Components

intermediate formation, intermediate terminated, assisted keto-enol tautomerisation, rate-determining step, proton transfer, dehydration, overall reactant used, unimolecular elimination by the conjugate base, intermediate collapse, overall product formed



Step 1.

Neutral Lys345 abstracts the proton alpha to the carbonyl group forming an oxyanion intermediate, which is stabilised by Lys396 and Mg(II) cations.



Step 2.

The oxyanion collapses, with concomitant double bond rearrangement, resulting in the elimination of the hydroxyl group, which abstracts a proton from the neutral Glu211.



Products.

The products of the reaction.

View Reaction Mechanisms in M-CSA

Reaction Parameters

• Kinetic Parameters

  Organism	KM Value [mM]	Substrate	Comment
  Klebsiella pneumoniae	23.5	phosphoenolpyruvate	in 50 mM imidazole-HCl buffer, pH 7.8, with 1 mM MgSO4, 0.4 M KCl
  Homo sapiens	48.5	phosphoenolpyruvate	in 50 mM imidazole-HCl buffer, pH 6.8, with 3 mM MgSO4, 0.4 M KCl
  Saccharolobus solfataricus	58.44	phosphoenolpyruvate	65°C, pH not specified in the publication
  Theileria annulata	350	2-phospho-D-glycerate	1.5 mM MgCl2 in 50 mM Tris/HCl (pH 7.4), at 25°C
  Plasmodium falciparum	520	2-phospho-D-glycerate	recombinant, dimeric enzyme

• Temperature

  Organism	Temperature Range	Comment
  Bacillus anthracis	25 - 45	at pH 6.8, recombinant protein
  Clostridioides difficile	25 - 80	80°C: 27% of maximal activity
  Pyrococcus furiosus	40 - 90	no activity at 25°C, activity increases from 40°C to 90°C, the highest assay temperature used

• pH

  Organism	pH Range	Comment
  Brucella abortus	5 - 10	activity range, recombinant enzyme
  Plasmodium falciparum	5 - 5.5	pH-dependent dissociation reveals that protonation of groups at the intersubunit interface is responsible for dissociation
  Macaca mulatta	6 - 9	pH 6.0: about 35% of maximal activity, pH 9.0: about 30% of maximal activity
  Trichinella spiralis	6 - 7.5
  Leuconostoc mesenteroides	6 - 8.3	catalytic activity above and below pH 6.8 remains constant at 30% of maximal activity

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Associated Proteins

Citations

• Characterization and epitope prediction of phosphopyruvate hydratase from Penaeus monodon (black tiger shrimp).
• cDNA cloning and expression analysis of a phosphopyruvate hydratase gene from the chinese shrimp Fenneropenaeus chinensis.
• Enolase: a metabolic checkpoint behind diverse exhaustion stages of CD8+ T cells in chronic HBV and HCV.
• Molecular cloning of kuruma shrimp Marsupenaeus japonicus phosphopyruvate hydratase and its role in infection by white spot syndrome virus and Vibrio alginolyticus
• Electrochemiluminescence resonance energy transfer between a Ru-ZnMOF self-enhanced luminophore and a double quencher ZnONF@PDA to detect NSE.
• Large-scale preparation of yeast strains expressing condensates derived from a glycolytic enzyme via controlled dissolved oxygen levels under hypoxia.
• Steroid-responsive ocular flutter with truncal ataxia and anti-amino-terminal of α-enolase antibody.
• High-Fat Mouse Model to Explore the Relationship between Abnormal Lipid Metabolism and Enolase in Pancreatic Cancer.
• Pyruvate kinase and phosphopyruvate hydratase as novel IgE reactive proteins in prawn.
• Treatment of the CRND8 mouse model for cerebral amyloid angiopathy, exhibited increased levels of neuron specific enolase in brain tissue following long-term treatment with a modified C5a receptor agonist, accompanied by improved cognitive function.
• Soluble Enolase 1 of Candida albicans and Aspergillus fumigatus Stimulates Human and Mouse B Cells and Monocytes.