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Enzymatic characterization and functional implication of two structurally different isocitrate dehydrogenases from Xylella fastidiosa.
Biotechnol Appl Biochem. 2018 Mar; 65(2):230-237.BA

Abstract

Isocitrate dehydrogenase (IDH) is a key enzyme at the critical junction between the tricarboxylic acid cycle and the glyoxylate cycle. Most bacteria have only one IDH, while a few contain two IDH isozymes. The coexistence of two different type IDHs in one organism was little known. Xylella fastidiosa is a nutritionally fastidious plant pathogen that contains two structurally different IDHs, an NAD+ -dependent homodimeric IDH (diXfIDH) and an NADP+ -dependent monomeric IDH (monoXfIDH). Kinetic characterization showed that diXfIDH displayed 206-fold preferences for NAD+ over NADP+ , while monoXfIDH showed 13,800-fold preferences for NADP+ over NAD+ . The putative coenzyme crucial amino acids (Asp-268, Ile-269, and Ala-275 in diXfIDH, and Lys-589, His-590, and Arg-601 in monoXfIDH) were studied by site-directed mutagenesis. The coenzyme specificities of the three diXfIDH mutants (D268K, D268K/I269Y, and D268K/I269Y/A275V) were switched successfully from NAD+ to NADP+ . Meanwhile, the mutant monoXfIDHs (H590L/R601L and K589T/H590L/R601L) greatly reduced the affinity for NADP+ , but failed to improve the ability to use NAD+ and had similar affinity to NADP+ and NAD+ . The biochemical properties of diXfIDH and monoXfIDH were investigated in detail. This study gives a further insight into the determinants of the coenzyme specificity in both monomeric and dimeric forms of IDHs.

Authors+Show Affiliations

The Research Center of Life Omics and Health and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, Anhui, People's Republic of China.The Research Center of Life Omics and Health and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, Anhui, People's Republic of China. Department of Biochemistry and Molecular Biology, Bengbu Medical College, Bengbu, People's Republic of China.The Research Center of Life Omics and Health and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, Anhui, People's Republic of China.The Research Center of Life Omics and Health and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, Anhui, People's Republic of China.The Research Center of Life Omics and Health and Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, Anhui, People's Republic of China.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

28220528

Citation

Lv, Peipei, et al. "Enzymatic Characterization and Functional Implication of Two Structurally Different Isocitrate Dehydrogenases From Xylella Fastidiosa." Biotechnology and Applied Biochemistry, vol. 65, no. 2, 2018, pp. 230-237.
Lv P, Tang W, Wang P, et al. Enzymatic characterization and functional implication of two structurally different isocitrate dehydrogenases from Xylella fastidiosa. Biotechnol Appl Biochem. 2018;65(2):230-237.
Lv, P., Tang, W., Wang, P., Cao, Z., & Zhu, G. (2018). Enzymatic characterization and functional implication of two structurally different isocitrate dehydrogenases from Xylella fastidiosa. Biotechnology and Applied Biochemistry, 65(2), 230-237. https://doi.org/10.1002/bab.1560
Lv P, et al. Enzymatic Characterization and Functional Implication of Two Structurally Different Isocitrate Dehydrogenases From Xylella Fastidiosa. Biotechnol Appl Biochem. 2018;65(2):230-237. PubMed PMID: 28220528.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Enzymatic characterization and functional implication of two structurally different isocitrate dehydrogenases from Xylella fastidiosa. AU - Lv,Peipei, AU - Tang,Wanggang, AU - Wang,Peng, AU - Cao,Zhengyu, AU - Zhu,Guoping, Y1 - 2017/05/11/ PY - 2015/12/09/received PY - 2017/02/16/accepted PY - 2017/2/22/pubmed PY - 2018/6/7/medline PY - 2017/2/22/entrez KW - Xylella fastidiosa KW - biochemical properties KW - coenzyme specificity KW - isocitrate dehydrogenase KW - site-directed mutagenesis SP - 230 EP - 237 JF - Biotechnology and applied biochemistry JO - Biotechnol. Appl. Biochem. VL - 65 IS - 2 N2 - Isocitrate dehydrogenase (IDH) is a key enzyme at the critical junction between the tricarboxylic acid cycle and the glyoxylate cycle. Most bacteria have only one IDH, while a few contain two IDH isozymes. The coexistence of two different type IDHs in one organism was little known. Xylella fastidiosa is a nutritionally fastidious plant pathogen that contains two structurally different IDHs, an NAD+ -dependent homodimeric IDH (diXfIDH) and an NADP+ -dependent monomeric IDH (monoXfIDH). Kinetic characterization showed that diXfIDH displayed 206-fold preferences for NAD+ over NADP+ , while monoXfIDH showed 13,800-fold preferences for NADP+ over NAD+ . The putative coenzyme crucial amino acids (Asp-268, Ile-269, and Ala-275 in diXfIDH, and Lys-589, His-590, and Arg-601 in monoXfIDH) were studied by site-directed mutagenesis. The coenzyme specificities of the three diXfIDH mutants (D268K, D268K/I269Y, and D268K/I269Y/A275V) were switched successfully from NAD+ to NADP+ . Meanwhile, the mutant monoXfIDHs (H590L/R601L and K589T/H590L/R601L) greatly reduced the affinity for NADP+ , but failed to improve the ability to use NAD+ and had similar affinity to NADP+ and NAD+ . The biochemical properties of diXfIDH and monoXfIDH were investigated in detail. This study gives a further insight into the determinants of the coenzyme specificity in both monomeric and dimeric forms of IDHs. SN - 1470-8744 UR - https://www.unboundmedicine.com/medline/citation/28220528/Enzymatic_characterization_and_functional_implication_of_two_structurally_different_isocitrate_dehydrogenases_from_Xylella_fastidiosa_ L2 - https://doi.org/10.1002/bab.1560 DB - PRIME DP - Unbound Medicine ER -