Isocitrate dehydrogenase (ICDH) (EC 126.96.36.199 and EC 188.8.131.52) is an enzyme that catalyzes the oxidative decarboxylation of isocitrate to 2-oxoglutarate. In prokaryotes, two types of phylogenetically unrelated ICDHs are known: monomeric and oligomeric. Prokaryotic oligomeric ICDH is evolutionarily related to isopropylmalate dehydrogenase (EC 184.108.40.206), tartrate dehydrogenase (EC 220.127.116.11), and homoisocitrate dehydrogenase (EC 18.104.22.168 and EC 22.214.171.1246); these enzymes constitute a group called NAD(P)-dependent β-hydroxyacid oxidative decarboxylases or NAD(P)-dependent β-decarboxylating dehydrogenases. The three-dimensional structures of Escherichia coli
ICDH (EcICDH), isopropylmalate dehydrogenase from Thermus thermophilus
, and homoisocitrate dehydrogenase from T. thermophilus
demonstrate that these enzymes share a common fold (11
). Prokaryotic ICDH in this group has previously been considered a homodimeric and an NAD(P)-dependent enzyme. However, due to increasing reports of NAD-dependent ICDH in bacteria (Acidithiobacillus
, and Streptococcus
) and archaea (Pyrococcus
) and of homotetrameric ICDH in bacteria (Methylococcus
), prokaryotic oligomeric ICDH is now recognized as an enzyme with various oligomeric states and coenzyme specificities.
Phylogenetic analyses of prokaryotic oligomeric ICDH indicate that this enzyme does not comprise a single lineage but can be divided into many subfamilies (21
). EcICDH is one of the best analyzed forms and belongs to a distinctive subfamily that also contains ICDH from archaea (Aeropyrum
, and Pyrococcus
) and Aquificales
). These enzymes can be considered a single lineage and can be categorized as EcICDH-type enzymes.
We have previously reported an EcICDH-type enzyme from an organism belonging to the order Aquificales
, Hydrogenobacter thermophilus
). The primary sequence of ICDH from H. thermophilus
(HtICDH) is 45.8% identical to that of EcICDH, although its enzymatic characteristics are quite different (3
). In particular, the physiological function of HtICDH is distinct from that of EcICDH. While EcICDH is involved in the tricarboxylic acid (TCA) cycle and catalyzes the oxidative decarboxylation of isocitrate, HtICDH is involved in the reductive TCA cycle and catalyzes the reduction of oxalosuccinate (2
) (Fig. ). Thus, differences in the reaction mechanism between these two enzymes were of great interest.
FIG. 1. Physiological roles of ICDH in E. coli and in H. thermophilus. (A) EcICDH is an enzyme involved in the TCA cycle and catalyzes the oxidative decarboxylation of isocitrate to form 2-oxoglutarate. (B) HtICDH is an enzyme involved in the reductive TCA cycle (more ...)
In this study, we analyzed the kinetic mechanism of the oxidation reaction catalyzed by HtICDH. As a result, we clearly demonstrate here that HtICDH is not a conventional decarboxylating ICDH but a novel nondecarboxylating ICDH. Furthermore, we suggest a possible hypothesis concerning the evolutionary history of the prokaryotic oligomeric ICDH where the oxalosuccinate-reducing enzyme is the ancestral form of the decarboxylating ICDHs.