These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: Distances among coenzyme and metal sites of NADP+-dependent isocitrate dehydrogenase using resonance energy transfer.
    Author: Bailey JM, Colman RF.
    Journal: Biochemistry; 1987 Jul 28; 26(15):4893-900. PubMed ID: 3663631.
    Abstract:
    When the substrate isocitrate-Mn2+ is present, the fluorescent nucleotide analogue 2-[(4-bromo-2,3-dioxobutyl)thio]-1,N6-ethenoadenosine 2',5'-bisphosphate (2-BDB-T epsilon A-2',5'-DP) reacts irreversibly with pig heart NADP+-specific isocitrate dehydrogenase at the coenzyme binding site on one subunit of the dimeric enzyme [Bailey, J. M., & Colman, R. F. (1985) Biochemistry 24, 5367-5377]. The modified enzyme, which retains partial activity, binds 1 mol of NADPH or 1 mol of the coenzyme analogue, reduced thionicotinamide adenine dinucleotide phosphate (TNADPH), per dimer. TNADPH quenches the fluorescence of enzyme-bound 2-BDB-T epsilon A-2',5'-DP with an efficiency of energy transfer of 9.8%. From this value and the spectral properties of the donor and acceptor chromophores, a distance of 32 A was calculated as the average distance between coenzyme sites on the two subunits. Isocitrate dehydrogenase activity requires a divalent metal ion, such as Mn2+, Co2+, or Ni2+. Co2+ and Ni2+ have absorption spectra that overlap the emission spectra of enzyme-bound 2-BDB-T epsilon A-2',5'-DP. In the presence of isocitrate, each of these two metal ions quenches the fluorescence of the enzyme-bound reagent with an efficiency of energy transfer of 28-29%. From this value and the spectral characteristics of the energy donor and acceptors, an average distance of 8.0 A was estimated between the metal-isocitrate site and the labeled coenzyme site. These distances have provided constraints in formulating a model of the spatial arrangement of active-site ligands on isocitrate dehydrogenase.
    [Abstract] [Full Text] [Related] [New Search]