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17O electron nuclear double resonance characterization of substrate binding to the [4Fe-4S]1+ cluster of reduced active aconitase.
J Biol Chem. 1986 Apr 15; 261(11):4840-6.JB

Abstract

To characterize the binding of substrate to aconitase, we have made 17O electron nuclear double resonance (ENDOR) measurements on reduced active ([4Fe-4S]1+) beef heart aconitase, both in H216O and H217O, in the presence of substrate and the inhibitors, tricarballylate, trans-aconitate, and 1-hydroxy-2-nitro-1, 3-propanedicarboxylate, referred to here as nitroisocitrate; the hydroxyl of the latter also was isotypically labeled with 17O. The hydroxyl oxygen of citrate and isocitrate is exchanged with solvent water by aconitase, but the hydroxyl of nitroisocitrate is not. Thus, the isotopic composition of nitroisocitrate can be chemically controlled, allowing direct identification of any 17O ENDOR signal associated with it. 17O ENDOR signals were observed from Hx17O (mean = 1 or 2) bound to the [4Fe-4S]1+ cluster in samples prepared with trans-aconitate and unlabeled nitroisocitrate. 17O-Labeled nitroisocitrate in H216O bound to the cluster showed a signal from the 17OH group; in H217O it showed 17O ENDOR resonances due to both Hx17O and 17OH of substrate. This result demonstrates that the cluster participates in substrate binding and can simultaneously coordinate the hydroxyl of a substrate (or analogue) and water (or hydroxyl). The sample with citrate in H217O showed only the Hx17O signal, although aconitase exchanges the hydroxyl of substrate with solvent water. The mechanism of action of aconitase is discussed in light of this observation. Comparison shows the ENDOR study to be in agreement with previous Mössbauer and EPR spectroscopic results.

Authors

Telser J
No affiliation info available
Emptage MH
No affiliation info available
Merkle H
No affiliation info available
Kennedy MC
No affiliation info available
Beinert H
No affiliation info available
Hoffman BM
No affiliation info available

MeSH

Aconitate HydrataseAconitic AcidAnimalsCattleCitratesCitric AcidElectron Spin Resonance SpectroscopyIronIsocitratesMyocardiumSulfurTricarboxylic Acids

Pub Type(s)

Comparative Study
Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

3007476

Citation

Telser, J, et al. "17O Electron Nuclear Double Resonance Characterization of Substrate Binding to the [4Fe-4S]1+ Cluster of Reduced Active Aconitase." The Journal of Biological Chemistry, vol. 261, no. 11, 1986, pp. 4840-6.
Telser J, Emptage MH, Merkle H, et al. 17O electron nuclear double resonance characterization of substrate binding to the [4Fe-4S]1+ cluster of reduced active aconitase. J Biol Chem. 1986;261(11):4840-6.
Telser, J., Emptage, M. H., Merkle, H., Kennedy, M. C., Beinert, H., & Hoffman, B. M. (1986). 17O electron nuclear double resonance characterization of substrate binding to the [4Fe-4S]1+ cluster of reduced active aconitase. The Journal of Biological Chemistry, 261(11), 4840-6.
Telser J, et al. 17O Electron Nuclear Double Resonance Characterization of Substrate Binding to the [4Fe-4S]1+ Cluster of Reduced Active Aconitase. J Biol Chem. 1986 Apr 15;261(11):4840-6. PubMed PMID: 3007476.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - 17O electron nuclear double resonance characterization of substrate binding to the [4Fe-4S]1+ cluster of reduced active aconitase. AU - Telser,J, AU - Emptage,M H, AU - Merkle,H, AU - Kennedy,M C, AU - Beinert,H, AU - Hoffman,B M, PY - 1986/4/15/pubmed PY - 1986/4/15/medline PY - 1986/4/15/entrez SP - 4840 EP - 6 JF - The Journal of biological chemistry JO - J Biol Chem VL - 261 IS - 11 N2 - To characterize the binding of substrate to aconitase, we have made 17O electron nuclear double resonance (ENDOR) measurements on reduced active ([4Fe-4S]1+) beef heart aconitase, both in H216O and H217O, in the presence of substrate and the inhibitors, tricarballylate, trans-aconitate, and 1-hydroxy-2-nitro-1, 3-propanedicarboxylate, referred to here as nitroisocitrate; the hydroxyl of the latter also was isotypically labeled with 17O. The hydroxyl oxygen of citrate and isocitrate is exchanged with solvent water by aconitase, but the hydroxyl of nitroisocitrate is not. Thus, the isotopic composition of nitroisocitrate can be chemically controlled, allowing direct identification of any 17O ENDOR signal associated with it. 17O ENDOR signals were observed from Hx17O (mean = 1 or 2) bound to the [4Fe-4S]1+ cluster in samples prepared with trans-aconitate and unlabeled nitroisocitrate. 17O-Labeled nitroisocitrate in H216O bound to the cluster showed a signal from the 17OH group; in H217O it showed 17O ENDOR resonances due to both Hx17O and 17OH of substrate. This result demonstrates that the cluster participates in substrate binding and can simultaneously coordinate the hydroxyl of a substrate (or analogue) and water (or hydroxyl). The sample with citrate in H217O showed only the Hx17O signal, although aconitase exchanges the hydroxyl of substrate with solvent water. The mechanism of action of aconitase is discussed in light of this observation. Comparison shows the ENDOR study to be in agreement with previous Mössbauer and EPR spectroscopic results. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/3007476/17O_electron_nuclear_double_resonance_characterization_of_substrate_binding_to_the_[4Fe_4S]1+_cluster_of_reduced_active_aconitase_ DB - PRIME DP - Unbound Medicine ER -