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. In humans, deficiency of ENO1 is linked to hereditary haemolytic anemia, while ENO3 deficiency is linked to glycogen storage disease type XIII. Isozymes In humans there are three subunits of enolase, α, β, and γ, each encoded by a separate gene that can combine to form five different isoenzymes: αα, αβ, αγ, ββ, and γγ. 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. Structure Enolase is a member of the large enolase superfamily. It has a molecular weight of 82,000-100,000 Daltons depending on the isoform. In human alpha enolase, the two subunits are antiparallel in orientation so that Glu20 of one subunit forms an ionic bond with Arg414 of the other subunit. Each subunit has two distinct domains. The smaller N-terminal domain consists of three α-helices and four β-sheets. The larger C-terminal domain starts with two β-sheets followed by two α-helices and ends with a barrel composed of alternating β-sheets and α-helices arranged so that the β-beta sheets are