title: Purification and characterization of cytosolic aconitase from beef liver and its relationship to the iron-responsive element binding protein.
abstract: In recent reports attention has been drawn to the extensive amino acid homology between pig heart, yeast, and Escherichia coli aconitases (EC 4.2.1.3) and the iron-responsive element binding protein (IRE-BP) of mammalian cells [Rouault, T. A., Stout, C. D., Kaptain, S., Harford, J. B. & Klausner, R. D. (1991) Cell 64, 881-883.; Hentze, M. W. & Argos, P. (1991) Nucleic Acids Res. 19, 1739-1740.; Prodromou, C., Artymiuk, P. J. & Guest, J. R. (1992) Eur. J. Biochem. 204, 599-609]. Iron-responsive elements (IREs) are stem-loop structures located in the untranslated regions of mRNAs. IRE-BP is required in the posttranscriptional regulation of ferritin mRNA translation and stabilization of transferrin receptor mRNA. In spite of substantial homology between the amino acid sequences of mammalian mitochondrial aconitase and IRE-BP, the mitochondrial protein does not bind IREs. However, there is a second aconitase, found only in the cytosol of mammalian tissues, that might serve as an IRE-BP. To test this possibility, we have prepared sufficient quantities of the heretofore poorly characterized beef liver cytosolic aconitase. This enzyme is isolated largely in its active [4Fe-4S] form and has a turnover number similar to that of mitochondrial aconitase. The EPR spectra of the two enzymes are markedly different. The amino acid composition, molecular weight, isoelectric point, and the sequences of six random peptides clearly show that these physicochemical and structural characteristics are identical to those of IRE-BP, and that c-aconitase is distinctly different from m-aconitase. In addition, both cytosolic aconitase and IRE-BP can have aconitase activity or function as IRE-BPs, as shown in the following paper and elsewhere [Zheng, L. Kennedy, M. C., Blondin, G. A., Beinert, H. & Zalkin, H. (1992) Arch. Biochem. Biophys., in press]. This leads us to the conclusion that cytosolic aconitase is IRE-BP.
authors.affiliation: Department of Biochemistry, Medical College of Wisconsin, Milwaukee 53226.
authors.forename: M C
authors.initials: MC
authors.lastname: Kennedy
authors.forename: L
authors.initials: L
authors.lastname: Mende-Mueller
authors.forename: G A
authors.initials: GA
authors.lastname: Blondin
authors.forename: H
authors.initials: H
authors.lastname: Beinert
mesh_terms: D000154:Aconitate Hydratase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002417:Cattle; D003600:Cytosol / Q000201:enzymology; D004578:Electron Spin Resonance Spectroscopy; D007501:Iron / Q000378:metabolism*; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins / Q000737:chemistry; D007526:Isoelectric Point; D008099:Liver / Q000201:enzymology*; D008930:Mitochondria, Liver / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry; D016601:RNA-Binding Proteins / Q000737:chemistry*; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D035925:Iron-Regulatory Proteins; D007506:Iron-Sulfur Proteins; D010446:Peptide Fragments; D016601:RNA-Binding Proteins; D007501:Iron; D000154:Aconitate Hydratase