ASP 241;GLU 266;ASN 295(Magnesium binding)
ASP 235;GLU 260;ASN 289(Magnesium binding)
The enolase superfamily: a general strategy for enzyme-catalyzed abstraction of the alpha-protons of carboxylic acids.
Evolution of enzymatic activities in the enolase superfamily: crystal structure of (D)-glucarate dehydratase from Pseudomonas putida.
Evolution of enzymatic activities in the enolase superfamily: partitioning of reactive intermediates by (D)-glucarate dehydratase from Pseudomonas putida.
Evolution of enzymatic activities in the enolase superfamily: crystallographic and mutagenesis studies of the reaction catalyzed by D-glucarate dehydratase from Escherichia coli.
Evolution of enzymatic activities in the enolase superfamily: identification of the general acid catalyst in the active site of D-glucarate dehydratase from Escherichia coli.
This enzyme belongs to the enolase superfamily. This enzyme catalyzes dehydration of either D-glucarate or L-idarate to form 5-Dehydro-4-deoxy-D-glucarate (KDG). According to the literature [2], [4] & [5], the catalytic reaction proceeds as follows: (A) Isomerization (change in the position of double-bond): (A1) A general base abstracts the alpha-proton from the C5 carbon, resulting in the formation of the enediolate anion intermediate. Here, His339 (of 1ec7) acts as the R-specific base for the D-glucarate substrate, whilst Lys207 acts as the S-specific base for the L-idarate (see [4] & [5]). (A2) The enediolate anion is stabilized by the manesium ion, which is coordinated by Asp235, Glu266 and Asn289. (B) Elimination of hydroxyl group from the substrate: (B1) A general acid eliminates the 4-OH group from the enediolate anion intermediate, by donating a proton to the group, resulting in the formation of an enol intermediate, in which a double bond is formed between the C4 and the C5 carbon atoms. Although the earlier paper [3] mentioned that Tyr150 may act as the general acid, more recent papers [4] & [5] proposed that His339 would act as the general acid. (B2) The enol intermediate might be stabilized by Tyr150, Lys205 and Asn237, as well as the magnesium ion (see [4]). The enol oxygen is stabilized by Tyr150, whereas the carboxylate is stabilized by Lys205, Asn237 and the magnesium ion. (C) Isomerization (change in the position of double-bond): (C0) The enol oxygen is stabilized by Tyr150, whereas the carboxylate is stabilized by Lys205, Asn237 and the magnesium ion. (C1) Another general acid donates a proton to the C4 carbon of the enol intermediate, leading to its tautomerization and to the final product, KDG. According to the paper [5], His339 acts as the second acid, as well.