Гликолиз ферменті: Енолазаның құрылымы мен қатысы

An enzyme involved in glycolysis

Phosphopyruvate hydratase, usually known as enolase, is a metalloenzyme that catalyses the conversion of 2 phosphoglycerate (2 PG) to phosphoenolpyruvate (PEP), the ninth and penultimate step of glycolysis. The chemical reaction is:

2 phospho D glycerate phosphoenolpyruvate + H2O

Phosphopyruvate hydratase belongs to the family of lyases, specifically the hydro lyases, which cleave carbon oxygen bonds. The systematic name of this enzyme is 2 phospho D glycerate hydro lyase (phosphoenolpyruvate forming). The reaction is reversible, depending on environmental concentrations of substrates. The optimum pH for the human enzyme is 6.5. Enolase is present in all tissues and organisms capable of glycolysis or fermentation. The enzyme was discovered by Lohmann and Meyerhof in 1934, and has since been isolated from a variety of sources including human muscle and erythrocytes. Three of these isoenzymes (all homodimers) are more commonly found in adult human cells than the others:

αα or non neuronal enolase (NNE). Also known as enolase 1. Found in a variety of tissues, including liver, brain, kidney, spleen, adipose. It is present at some level in all normal human cells. ββ or muscle specific enolase (MSE). Also known as enolase 3. This enzyme is largely restricted to muscle where it is present at very high levels in muscle. γγ or neuron specific enolase (NSE). Also known as enolase 2. Expressed at very high levels in neurons and neural tissues, where it can account for as much as 3% of total soluble protein. It is expressed at much lower levels in most mammalian cells. When present in the same cell, different isozymes readily form heterodimers.

Enolase is a member of the large enolase superfamily. It has a molecular weight of 82,000–100,000 daltons depending on the isoform.

Using isotopic probes, the overall mechanism for converting 2 PG to PEP is proposed to be an E1cB elimination reaction involving a carbanion intermediate. The following detailed mechanism is based on studies of crystal structure and kinetics. When the substrate, 2 phosphoglycerate, binds to α enolase, its carboxyl group coordinates with two magnesium ion cofactors in the active site. This stabilizes the negative charge on the deprotonated oxygen while increasing the acidity of the alpha hydrogen. Enolase's Lys345 deprotonates the alpha hydrogen, and the resulting negative charge is stabilized by resonance to the carboxylate oxygen and by the magnesium ion cofactors. Following the creation of the carbanion intermediate, the hydroxide on C3 is eliminated as water with the help of Glu211, and PEP is formed. Additionally, conformational changes occur within the enzyme that aid catalysis. In human α enolase, the substrate is rotated into position upon binding to the enzyme due to interactions with the two catalytic magnesium ions, Gln167, and Lys396. Movements of loops Ser36 to His43, Ser158 to Gly162, and Asp255 to Asn256 allow Ser39 to coordinate with Mg2+ and close off the active site. In addition to coordination with the catalytic magnesium ions, the pKa of the substrate's alpha hydrogen is also lowered due to protonation of the phosphoryl group by His159 and its proximity to Arg374. Arg374 also causes Lys345 in the active site to become deprotonated, which primes Lys345 for its role in the mechanism.

In recent medical experiments, enolase concentrations have been sampled in an attempt to diagnose certain conditions and their severity. For example, higher concentrations of enolase in cerebrospinal fluid more strongly correlated to low grade astrocytoma than did other enzymes tested (aldolase, pyruvate kinase, creatine kinase, and lactate dehydrogenase). The same study showed that the fastest rate of tumor growth occurred in patients with the highest levels of CSF enolase. Increased levels of enolase have also been identified in patients who have suffered a recent myocardial infarction or cerebrovascular accident. It has been inferred that levels of CSF neuron specific enolase, serum NSE, and creatine kinase (type BB) are indicative in the prognostic assessment of cardiac arrest victims. Other studies have focused on the prognostic value of NSE values in cerebrovascular accident victims. Autoantibodies to alpha enolase are associated with rheumatoid arthritis and the rare syndrome called Hashimoto's encephalopathy.

Small molecule inhibitors of enolase have been synthesized as chemical probes (substrate analogues) of the catalytic mechanism of the enzyme and more recently, have been investigated as potential treatments for cancer and infectious diseases. Most inhibitors have metal chelating properties and bind to enzyme by interactions with the structural Magnesium Atom Mg(A). The most potent of these is phosphonoacetohydroxamate, and more recently, as an anti cancer agent, specifically, in glioblastoma that are enolase deficient due to homozygous deletion of the ENO1 gene as part of the 1p36 tumor suppressor locus (synthetic lethality). A natural product phosphonate antibiotic, SF2312 (CAS 107729 45 3), which is active against gram positive and negative bacteria especially under anaerobic conditions, is a high potency inhibitor of Enolase that binds in manner similar to phoshphonoacetohydroxamate SF2312 inhibits Enolase activity in both eukaryotic and prokaryotic origin, reflecting the strong evolutionary conservation of Enolase and the ancient origin of the glycolysis pathway. SF2312 is a chiral molecule with only the 3S enantiomer showing Enolase inhibitory activity and biological activity against bacteria. More recently, a derivative of SF2312, termed HEX, and a prodrug thereoff, POMHEX, were shown to exert anti neoplastic activity against ENO1 deleted glioma in a pre clinical intracranial orthotopic mouse model. An allosteric binder, ENOblock ENOblock was found to alter the cellular localization of enolase, influencing its secondary, non glycolytic functions, such as transcription regulation. Subsequent analysis using a commercial assay also indicated that ENOblock can inhibit enolase activity in biological contexts, such as cells and animal tissues. Active site transition state analogue Enolase inhibitors have been explored pre clinically for the treatment of various microbial pathogens, as well as in precision oncology for tumors with 1p36 homozygous deletions, that lack ENO1. Fluoride is a known competitor of enolase's substrate 2 PG. Fluoride can form a complex with magnesium and phosphate, which binds in the active site instead of 2 PG. The Enolase inhibitory activity of Fluoride anion may contribute to the anti cavity effect of fluoride toothpaste, by limiting lactic acid (a product of glycolysis, which requires Enolase) production.