Skip to main page content
U.S. flag
See More

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2008 Jul;42(7):618-24.
doi: 10.1080/10715760802209639.

An SOD mimic protects NADP+-dependent isocitrate dehydrogenase against oxidative inactivation

Ines Batinic-Haberle  1 , Ludmil T Benov
Affiliations

An SOD mimic protects NADP+-dependent isocitrate dehydrogenase against oxidative inactivation

Ines Batinic-Haberle et al. Free Radic Res. 2008 Jul.

Abstract

The isocitrate dehydrogenases (ICDs) catalyse the oxidative decarboxylation of isocitrate to alpha-ketoglutarate and can use either NAD(+) or NADP(+) as a cofactor. Recent studies demonstrate that the NADP(+)-dependent isocitrate dehydrogenase, as a source of electrons for cellular antioxidants, is important for protection against oxidative damage. ICD, however, is susceptible to oxidative inactivation, which in turn compromises cellular antioxidant defense. This study investigates the effect of a superoxide dismutase (SOD) mimic, MnTM-2-PyP(5+), on the inactivation of NADP(+)-dependent ICD in SOD-deficient Escherichia coli and in diabetic rats. The findings show that E. coli ICD is inactivated by superoxide, but the inactivated enzyme is replaced by de novo protein synthesis. Statistically significant decrease of ICD activity was found in the hearts of diabetic rats. MnTM-2-PyP(5+) protected ICD in both models.

PubMed Disclaimer

Conflict of interest statement

Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.

Figures

Figure 1
Figure 1
MnTM-2-PyP5+, Mn(III) tetrakis(N-methylpyridinium-2-yl)porphyrin.
Figure 2
Figure 2
Inactivation of ICD by enzymatically generated superoxide. E. coli cell extracts (~1.0 mg protein/ml) were incubated at 37°C in the presence of 500 µm hypoxanthine and xanthine oxidase to give an initial superoxide generation rate of 5 µm/min. Experiments were repeated three times with 3–5 replicates. Means±SE are presented. Line 1, parental; Line 2, cell extract of sodAsodB cells grown with 20 µm of MnTM-2-PyP; Line 3, sodAsodB cell extracts plus 20 µm MnTM-2-PyP; Line 4, sodAsodB cell extracts.
Figure 3
Figure 3
Aconitase activity in parental and SOD-deficient cells grown with or without MnTM-2-PyP. E. coli cultures were grown to a density of A600nm 0.7–0.8 with or without 20 µm of MnTM-2-PyP. The cells were washed, disrupted by sonication and aconitase was assayed. The assay mixture contained 50 mm Tris-Cl buffer, pH 7.4, 30 mm sodium citrate, 0.5 mm MnCl2, 0.2 mm NADP+, 2.0 units/ml of isocitrate dehydrogenase and cell extract. Results are presented as a percentage of the total aconitase activity obtained after reactivation of the enzyme. Experiments were repeated three times with 3–5 replicates. Bars represent means±SE.
Figure 4
Figure 4
Isocitrate dehydrogenase activity in exponentially growing E. coli cultures. E. coli parental and SOD-deficient strains were grown in LB medium to the indicated densities. The cells were washed, disrupted by sonication and ICD activity was measured in 1.0 ml reaction mixture contained 33 mm Tris-EDTA buffer (pH 7.4), 1.33 mm MnCl2, 1.3 mm DL-Isocitrate and 0.1 mm NADP+. Experiments were repeated three times with 3–5 replicates. Bars represent means±SE.
Figure 5
Figure 5
Effect of spectinomycin and paraquat on isocitrate dehydrogenase activity. E. coli parental and SOD-deficient strains were grown in LB medium to a density of A600nm 0.6. Spectinomycin was added to 500 µm, followed 15 min later by addition of paraquat to 50 µm. One hour after the addition of paraquat the cells were washed, disrupted by sonication and ICD activity was measured. Experiments were repeated three times with 3–5 replicates. Bars represent means±SE.
Figure 6
Figure 6
Activities of isocitrate dehydrogenase, aconitase and fumarase in rat hearts. The enzyme activities were measured in heart homogenates of control, non-diabetic MnTM-2-PyP-treated, diabetic and diabetic MnTM-2-PyP-treated rats. This experiment was repeated twice with 7–12 animals per group. Bars represent means±SE. * p < 0.05 compared to non-treated group.
See this image and copyright information in PMC

References

    1. Xu X, Zhao J, Xu Z, Peng B, Huang Q, Arnold E, Ding J. Structures of human cytosolic NADP-dependent isocitrate dehydrogenase reveal a novel self-regulatory mechanism of activity. J Biol Chem. 2004;279:33946–33957. - PubMed
    1. Haselbeck RJ, McAlister-Henn L. Function and expression of yeast mitochondrial NAD- and NADP-specific isocitrate dehydrogenases. J Biol Chem. 1993;268:12116–12122. - PubMed
    1. Jennings GT, Sechi S, Stevenson PM, Tuckey RC, Parmelee D, McAlister-Henn L. Cytosolic NADP(+)-dependent isocitrate dehydrogenase. Isolation of rat cDNA and study of tissue-specific and developmental expression of mRNA. J Biol Chem. 1994;269:23128–23134. - PubMed
    1. Jo SH, Son MK, Koh HJ, Lee SM, Song IH, Kim YO, Lee YS, Jeong KS, Kim WB, Park JW, Song BJ, Huh TL. Control of mitochondrial redox balance and cellular defense against oxidative damage by mitochondrial NADP+-dependent isocitrate dehydrogenase. J Biol Chem. 2001;276:16168–16176. - PubMed
    1. Lee SM, Koh HJ, Park DC, Song BJ, Huh TL, Park JW. Cytosolic NADP(+)-dependent isocitrate dehydrogenase status modulates oxidative damage to cells. Free Radic Biol Med. 2002;32:1185–1196. - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources