Mode of substrate carboxyl binding to the (4Fe-4S)/sup +/ cluster of reduced aconitase as studied by /sup 17/O and /sup 13/C electron-nuclear double resonance spectroscopy

Journal Article · Tue Dec 01 04:00:00 EST 1987 · Proc. Natl. Acad. Sci. U.S.A.; (United States)

DOI:https://doi.org/10.1073/pnas.84.24.8854· OSTI ID:6351309

Kennedy, M C; Werst, M; Telser, J; Emptage, M H; Beinert, H; Hoffman, B M

The active form of aconitase has a diamagnetic (4Fe-4S)/sup 2 +/ cluster. A specific iron ion (Fe/sub a/, which is lost during inactivation) is the binding site for substrate, as shown by Moessbauer spectroscopy. The authors have studied the mode of substrate and analogue binding at equilibrium to the paramagnetic (4Fe-4S)/sup +/ cluster of the reduced active form by /sup 17/O and /sup 13/C electron-nuclear double resonance spectroscopy with specifically labeled substrates. The data show that with substrate, only the carboxyl at C-2 of the propane backbone is strongly bound in addition to H/sub 2/O or OH/sup -/ (H/sub x/O) from the solvent, whereas in an isocitrate analogue that has a nitro group at C-2, the carboxyl and hydroxyl at C-1 are bound along with solvent H/sub x/O. They conclude from these data that, on addition of any one of the three substrates, cis-aconitate is the predominant species bound to Fe/sub a/ of the cluster along with solvent H/sub x/O and that cis-aconitate is bound in the citrate mode (carboxyl at C-2). The results with the nitro analogue show that the enzyme can also bind a substrate-like ligand to the cluster in the alternative isocitrate mode (carboxyl at C-1), as is implicit in models proposed for the aconitase reaction.

• Research Organization: Medical College of Wisconsin, Milwaukee (USA)
• OSTI ID: 6351309
• Journal Information: Proc. Natl. Acad. Sci. U.S.A.; (United States), Journal Name: Proc. Natl. Acad. Sci. U.S.A.; (United States) Vol. 84:24; ISSN PNASA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
BIOCHEMISTRY
CARBON 13
CARBON ISOTOPES
CARBON-OXYGEN LYASES
CARBOXYLIC ACID SALTS
CHEMISTRY
CITRATES
ELECTRON SPECTRA
ENDOR
ENZYMES
EVEN-ODD NUCLEI
ISOTOPES
LABELLING
LIGANDS
LIGHT NUCLEI
LYASES
MAGNETIC RESONANCE
NUCLEI
OXYGEN 17
OXYGEN ISOTOPES
RESONANCE
SPECTRA
STABLE ISOTOPES
STEREOCHEMISTRY
SUBSTRATES