Abstract
Isocitrate dehydrogenase (IDH) catalyzes the reversible conversion between isocitrate and 2-oxoglutarate accompanied by decarboxylation/carboxylation and oxidoreduction of NAD(P)+ cofactor. While this enzyme has been well studied as a catabolic enzyme in the tricarboxylic acid (TCA) cycle, here we have characterized NADP-dependent IDH from Chlorobium limicola, a green sulfur bacterium that fixes CO2 through the reductive tricarboxylic acid (RTCA) cycle, focusing on the CO2-fixation ability of the enzyme. The gene encoding Cl-IDH consisted of 2226 bp, corresponding to a polypeptide of 742 amino acid residues. The primary structure and the size of the recombinant protein indicated that Cl-IDH was a monomeric enzyme of 80 kDa distinct from the dimeric NADP-dependent IDHs predominantly found in bacteria or eukaryotic mitochondria. Apparent Michaelis constants for isocitrate (45 ± 13 μM) and NADP+ (27 ± 10 μM) were much smaller than those for 2-oxoglutarate (1.1 ± 0.5 mM) and CO2 (1.3 ± 0.3 mM). No significant differences in kinetic properties were observed between Cl-IDH and the dimeric, NADP-dependent IDH from Saccharomyces cerevisiae (Sc-IDH) at the optimum pH of each enzyme. However, in contrast to the 20% activity of Sc-IDH toward carboxylation as compared with that toward decarboxylation at pH 7.0, the activities of Cl-IDH for both directions were almost equivalent at this pH, suggesting a more favorable property of Cl-IDH than Sc-IDH as a CO2-fixation enzyme under physiological pH. Furthermore, we found that among various intermediates, oxaloacetate was a competitive inhibitor (Ki = 0.35 ± 0.04 mM) for 2-oxoglutarate in the carboxylation reaction by Cl-IDH, a feature not found in Sc-IDH.
| Original language | English |
|---|---|
| Pages (from-to) | 1926-1931 |
| Number of pages | 6 |
| Journal | European Journal of Biochemistry |
| Volume | 269 |
| Issue number | 7 |
| DOIs | |
| Publication status | Published - 2002 |
| Externally published | Yes |
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Keywords
- CO-fixing enzyme
- Isocitrate dehydrogenase
- Reductive tricarboxylic acid cycle
ASJC Scopus subject areas
- Biochemistry
Cite this
Characterization of isocitrate dehydrogenase from the green sulfur bacterium chlorobium limicola : A carbon dioxide-fixing enzyme in the reductive tricarboxylic acid cycle. / Kanao, Tadayoshi; Kawamura, Mineko; Fukui, Toshiaki; Atomi, Haruyuki; Imanaka, Tadayuki.
In: European Journal of Biochemistry, Vol. 269, No. 7, 2002, p. 1926-1931.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Characterization of isocitrate dehydrogenase from the green sulfur bacterium chlorobium limicola
T2 - A carbon dioxide-fixing enzyme in the reductive tricarboxylic acid cycle
AU - Kanao, Tadayoshi
AU - Kawamura, Mineko
AU - Fukui, Toshiaki
AU - Atomi, Haruyuki
AU - Imanaka, Tadayuki
PY - 2002
Y1 - 2002
N2 - Isocitrate dehydrogenase (IDH) catalyzes the reversible conversion between isocitrate and 2-oxoglutarate accompanied by decarboxylation/carboxylation and oxidoreduction of NAD(P)+ cofactor. While this enzyme has been well studied as a catabolic enzyme in the tricarboxylic acid (TCA) cycle, here we have characterized NADP-dependent IDH from Chlorobium limicola, a green sulfur bacterium that fixes CO2 through the reductive tricarboxylic acid (RTCA) cycle, focusing on the CO2-fixation ability of the enzyme. The gene encoding Cl-IDH consisted of 2226 bp, corresponding to a polypeptide of 742 amino acid residues. The primary structure and the size of the recombinant protein indicated that Cl-IDH was a monomeric enzyme of 80 kDa distinct from the dimeric NADP-dependent IDHs predominantly found in bacteria or eukaryotic mitochondria. Apparent Michaelis constants for isocitrate (45 ± 13 μM) and NADP+ (27 ± 10 μM) were much smaller than those for 2-oxoglutarate (1.1 ± 0.5 mM) and CO2 (1.3 ± 0.3 mM). No significant differences in kinetic properties were observed between Cl-IDH and the dimeric, NADP-dependent IDH from Saccharomyces cerevisiae (Sc-IDH) at the optimum pH of each enzyme. However, in contrast to the 20% activity of Sc-IDH toward carboxylation as compared with that toward decarboxylation at pH 7.0, the activities of Cl-IDH for both directions were almost equivalent at this pH, suggesting a more favorable property of Cl-IDH than Sc-IDH as a CO2-fixation enzyme under physiological pH. Furthermore, we found that among various intermediates, oxaloacetate was a competitive inhibitor (Ki = 0.35 ± 0.04 mM) for 2-oxoglutarate in the carboxylation reaction by Cl-IDH, a feature not found in Sc-IDH.
AB - Isocitrate dehydrogenase (IDH) catalyzes the reversible conversion between isocitrate and 2-oxoglutarate accompanied by decarboxylation/carboxylation and oxidoreduction of NAD(P)+ cofactor. While this enzyme has been well studied as a catabolic enzyme in the tricarboxylic acid (TCA) cycle, here we have characterized NADP-dependent IDH from Chlorobium limicola, a green sulfur bacterium that fixes CO2 through the reductive tricarboxylic acid (RTCA) cycle, focusing on the CO2-fixation ability of the enzyme. The gene encoding Cl-IDH consisted of 2226 bp, corresponding to a polypeptide of 742 amino acid residues. The primary structure and the size of the recombinant protein indicated that Cl-IDH was a monomeric enzyme of 80 kDa distinct from the dimeric NADP-dependent IDHs predominantly found in bacteria or eukaryotic mitochondria. Apparent Michaelis constants for isocitrate (45 ± 13 μM) and NADP+ (27 ± 10 μM) were much smaller than those for 2-oxoglutarate (1.1 ± 0.5 mM) and CO2 (1.3 ± 0.3 mM). No significant differences in kinetic properties were observed between Cl-IDH and the dimeric, NADP-dependent IDH from Saccharomyces cerevisiae (Sc-IDH) at the optimum pH of each enzyme. However, in contrast to the 20% activity of Sc-IDH toward carboxylation as compared with that toward decarboxylation at pH 7.0, the activities of Cl-IDH for both directions were almost equivalent at this pH, suggesting a more favorable property of Cl-IDH than Sc-IDH as a CO2-fixation enzyme under physiological pH. Furthermore, we found that among various intermediates, oxaloacetate was a competitive inhibitor (Ki = 0.35 ± 0.04 mM) for 2-oxoglutarate in the carboxylation reaction by Cl-IDH, a feature not found in Sc-IDH.
KW - CO-fixing enzyme
KW - Isocitrate dehydrogenase
KW - Reductive tricarboxylic acid cycle
UR - http://www.scopus.com/inward/record.url?scp=0036230979&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0036230979&partnerID=8YFLogxK
U2 - 10.1046/j.1432-1327.2002.02849.x
DO - 10.1046/j.1432-1327.2002.02849.x
M3 - Article
C2 - 11952794
AN - SCOPUS:0036230979
VL - 269
SP - 1926
EP - 1931
JO - FEBS Journal
JF - FEBS Journal
SN - 1742-464X
IS - 7
ER -