ENO1 (Enolase 1, (alpha))

Note	Alpha-Enolase (ENO1, alpha enolase, non-neuronal enolase) is one of the three enolase enzymes, expressed in a wide variety of tissues. The other two enolase genes, ENO2 and ENO3, encode gamma (neuron-specific) and beta (muscle-specific) enolase, respectively. The active enolase enzymes exist as homodimers of non-covalently bound subunits. Each alpha, beta or gamma subunit is encoded by separate genes. The genomic organisation of ENO1 gene is identical with that of human gamma-enolase gene. All the coding exons have exactly the same length and introns occur at analogous positions.
	Structure of ENO1 mRNA ; note that nucleotides number (nt), exon positions (1-12) and two translation initiation sites (ATG) are labelled.
Description	The ENO1 gene consists of 12 exons distributed over 17718 bp of genomic DNA. Single alpha-enolase transcript contains two translation initiation positions and encodes two structurally and functionally distinct proteins, alpha-enolase enzyme and MYC promotor-binding protein (MBP-1).
Transcription	Transcription start sites of ENO1 gene are heterogeneous and spread over 38-bp region located at 116 bp upstream from the initiation codon ATG. These multiple start sites of transcription in ENO1 gene are consistent with lack of a canonical TATA box, usually found at the position 19-27 bp upstream of the cap sites. It is worth to notice that promoter of ENO1 gene contains two perfect Myc-Max binding motifs CACGTC. Other regulatory sites found in the 5'-flanking region of ENO1 gene include AP1 (T[T/G]AGTCA), AP2 (CCCCAGGC), AP3 (GGGTGTGGAAAG), AP4 (CAGCTGTGG), AP5 (CTGTGGAATG), ATF/CREB ([T/G][A/T]CGTCA), C2 (CATGTG), CTF/NF1 (TGGCTNNNAGCCAA), E2AE-C beta (TGGGAATT), E2F (TTTCGCGC), E4TF1 (GGAAGTG), EF-C (GTTGCNNGGCAAC), MLTF/USF (GGTCACGTGGCC), Ig octamer (ATTTGCAT), PEA2 (GACCGCA), SP1 (GGGCGG), CACCC (may function as CAAT boxes) and viral core (GTGG[A/T][A/T][A/T]G).
Pseudogene	A pseudogene has been identified that is located on the other arm of the same chromosome (provided by RefSeq).

	Structure of ENO1/MBP-1 protein; N and C termini, and amino acid (aa) positions are labelled.
Description	Alpha-Enolase (ENO1), like two other isoenzymes (gamma-ENO2 and beta-ENO3), is made up of two identical (homodimer), non-covalently bound alpha alpha subunits; alpha-Enolase is resolved during 1D-PAGE as a protein with molecular weight of about 48kDa (434 amino acids). It was demonstrated that in brain and neurons, specific enolases may exist as heterodimers, such as alpha alpha, alpha beta, beta beta, alpha gamma and gamma gamma. The proportions of isoenzymes alpha alpha, alpha beta and beta beta change in heart and muscle during embryonic development. In both mentioned tissues, isoform alpha was found predominantly in fetus. In adult heart this subunit is replaced by types alpha beta and beta beta, and in muscle by type beta beta. In human adult brain tissues, apha-type and gamma-type enolase subunits are present at similar concentrations. Two identical subunits of alpha-Enolase facilitate each other in an antiparallel fashion. Each subunit is made up of two distinct domains: N-terminal domain, consisting of three beta-sheets and four alpha-helices (beta3 alpha4 topology), and larger C-terminal domain with eightfold alpha beta barrel structure with beta beta alpha alpha(beta alpha)6 topology. This domain contains two beta-sheets at the beginning, followed by two alpha-helices and ends with a barrel made up of alternating beta-sheets and alpha-helices (beta - sheets are surrounded by the alpha - helices). The N-terminal of one subunit contacts the C-terminal of the second in such way, that glutamic acid at position 20 (Glu20) forms an ionic pair with arginine at position 414 (Arg414). Alternatively translated product of ENO1 gene, called MBP-1 (MYC promotor-binding protein), is expressed as a 37kDa (338 amino acids) protein and does not posses the enolase enzyme activity.
Expression	Apha-Enolase is widely expressed in variety of tissues including liver, brain, kidney, spleen, adipose as well as thyroid. In comparison with gamma-type subunit found only in neurons, type alpha subunit was also detected in astrocytes, ependymal cells, capillary endothelial cells, Schwann cells and arachnoidal endothelial cells.
Localisation	Alpha-Enolase is most abundantly found in cytoplasm and also on the cell surface. MBP-1 is localised in the nucleus.
Function	Enolase enzymes (2-phospho-D-glycerate hydrolases) catalyse the dehydration of 2-phospho-D-glycerate (PGA) to phosphoenolopyruvate (PEP) in Emden Mayerhoff-Parnas glycolytic pathway (catabolic direction). In anabolic pathway (reverse reaction) during gluconeogenesis, the same enzyme catalyses hydration of PEP to PGA (hence it is called phosphopyruvate hydratase). Metal ions are cofactors impairing the increase of enolase activity; hence it is also called metal-activated metalloenzyme. Magnesium is a natural cofactor causing the highest activity. The relative activation strength profile of metal ions involved in the enzyme activity appears in the following rank of order Mg2+ > Zn2+ > Mn2+ > Fe(II)2+ > Cd2+ > Co2+, Ni2+, Sm3+, Tb3+ and most other divalent metal ions. In reaction catalyzed by enolases, the alpha-proton from a carbon adjacent to a carboxylate group of PGA, is abstracted, and PGA is conversed to enolate anion intermediate. This intermediate is further processed in a variety of chemical reactions, including racemization, cycloisomerization and beta-elimination of either water or ammonia. The smaller product of ENO1 gene, MBP-1, is known as c-myc binding protein and negative regulator of its expression. C-myc is a DNA-binding phosphoprotein and a key regulator of cell behaviour. Many of c-myc targeting pathways are deregulated in cancer cells and contribute to its enhanced expression. There are four c-myc promoters, designated as P0, P1, P2 and P3, although in normal and cancer cells most mRNAs initiate at the P2 promoter. MBP-1 binds in a region +123 to +153 relative to the c-myc P2 promoter and probably by preventing the formation of a transcription initiation complex, decrease c-myc promoter activity. Hence MBP-1 is considered as tumor suppressor. ENO1 protein was also found as a structural component of the eye lenses and was designated as tau-crystalin. ENO1 enzyme and tau-crystalin are the products of the same gene. Tau-crystalins are the major components of vertebrate lens. These proteins are mainly found in monomeric form with a low enzymatic activity, while the active ENO1 enzyme exists as a dimer. Irrespective of ENO1 enzyme activity, its significant presence in eye lens (23% of the total protein of the lens) clearly indicate ENO1 structural role in lens and cataracts. In hypoxic conditions elevated ENO1 levels may provide protection to the cells by increasing anaerobic metabolism.
	Reaction catalyzed by Enolase.
Homology	Currently, amino acid sequences of more than 50 enolase enzymes are known. The five residues that participate in catalytic activity of this enzyme are highly conserved throughout evolution. Studies in vitro revealed that mutant enolase enzymes that differs at either positions Glu168, Glu211, Lys345, Lys396 or His159, demonstrated dramatically decreased activity level. An integral and conserved part of enolases are two Mg2+ ions that participate in conformational changes of the active site of enolase and enable binding of a substrate or its analogues.

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