Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:O14944 (EPR)
 13,097 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have determined the formal potential values for each electron transfer to electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO), in order to further characterize the thermodynamics of electron transport from various acyl-CoA thioesters to the mitochondrial ubiquinone pool. ETF-QO contains one [4Fe-4S]2+,1+ cluster and one FAD prosthetic group. A preliminary visible-spectroelectrochemical titration showed that the two redox centers were reduced almost simultaneously. Since the visible spectra of the chromophores overlap, it was not possible to resolve the formal potential value for each electron transfer to the protein using this method. Accordingly, an EPR-spectroelectrochemical cell was designed so that each formal potential value could be resolved by EPR quantitation of the flavin semiquinone and the reduced iron-sulfur cluster during the titration. The formal potential values for electron transfer to ETF-ubiquinone oxidoreductase at pH 7.5 and 4 degrees C were E1 degrees' = +0.028 V and E2 degrees' = -0.006 V for the first and second electron transfers, respectively, to the FAD and E degrees' = +0.047 V for the iron-sulfur cluster. The thermodynamics of electron transport from the acyl-CoA substrates of beta-oxidation to the mitochondrial electron transport chain have been fully resolved with completion of this work. The results are discussed in terms of their significance to the overall electron transport process from beta-oxidation.
 ...
PMID:Redox properties of electron-transfer flavoprotein ubiquinone oxidoreductase as determined by EPR-spectroelectrochemistry. 133 70

The clinical and biochemical phenotype of glutaric acidaemia type II (GAII) has led to the suggestion that the defect in the disorder affects electron transfer from primary FAD-containing dehydrogenases into the respiratory chain. Two proteins are involved in this process, i.e. electron transfer flavoprotein (ETF) and ETF dehydrogenase, an iron--sulphur flavoprotein with a distinctive EPR signal. Reliable catalytic assays for these proteins are not available, but both proteins have been purified and antisera against them prepared in rabbits. SDS-PAG electrophoresis of liver mitochondrial membranes from a GAII infant with congenital anomalies, locating ETF dehydrogenase with specific antiserum, showed no cross-reactive material. EPR of the same membranes showed a marked decrease in the ETF dehydrogenase signal. These results suggest that the defect in GAII in some patients is indeed in electron transport, and specifically in ETF dehydrogenase.
 ...
PMID:Glutaric acidaemia type II (multiple acyl-CoA dehydrogenation deficiency). 643 42

Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) is an iron-sulphur flavoprotein and a component of an electron-transfer system that links 10 different mitochondrial flavoprotein dehydrogenases to the mitochondrial bc1 complex via electron transfer flavoprotein (ETF) and ubiquinone. ETF-QO is an integral membrane protein, and the primary sequences of human and porcine ETF-QO were deduced from the sequences of the cloned cDNAs. We have expressed human ETF-QO in Sf9 insect cells using a baculovirus vector. The cDNA encoding the entire protein, including the mitochondrial targeting sequence, was present in the vector. We isolated a membrane-bound form of the enzyme that has a molecular mass identical with that of the mature porcine protein as determined by SDS/PAGE and has an N-terminal sequence that is identical with that predicted for the mature holoenzyme. These data suggest that the heterologously expressed ETF-QO is targeted to mitochondria and processed to the mature, catalytically active form. The detergent-solubilized protein was purified by ion-exchange and hydroxyapatite chromatography. Absorption and EPR spectroscopy and redox titrations are consistent with the presence of flavin and iron-sulphur centres that are very similar to those in the equivalent porcine and bovine proteins. Additionally, the redox potentials of the two prosthetic groups appear similar to those of the other eukaryotic ETF-QO proteins. The steady-state kinetic constants of human ETF-QO were determined with ubiquinone homologues, a ubiquinone analogue, and with human wild-type ETF and a Paracoccus-human chimaeric ETF as varied substrates. The results demonstrate that this expression system provides sufficient amounts of human ETF-QO to enable crystallization and mechanistic investigations of the iron-sulphur flavoprotein.
 ...
PMID:Expression of human electron transfer flavoprotein-ubiquinone oxidoreductase from a baculovirus vector: kinetic and spectral characterization of the human protein. 1204 29

Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) accepts electrons from electron transfer flavoprotein (ETF) and reduces ubiquinone from the ubiquinone pool. It contains one [4Fe-4S] (2+,1+) and one FAD, which are diamagnetic in the isolated oxidized enzyme and can be reduced to paramagnetic forms by enzymatic donors or dithionite. In the porcine protein, threonine 367 is hydrogen bonded to N1 and O2 of the flavin ring of the FAD. The analogous site in Rhodobacter sphaeroides ETF-QO is asparagine 338. Mutations N338T and N338A were introduced into the R. sphaeroides protein by site-directed mutagenesis to determine the impact of hydrogen bonding at this site on redox potentials and activity. The mutations did not alter the optical spectra, EPR g-values, spin-lattice relaxation rates, or the [4Fe-4S] (2+,1+) to FAD point-dipole interspin distances. The mutations had no impact on the reduction potential for the iron-sulfur cluster, which was monitored by changes in the continuous wave EPR signals of the [4Fe-4S] (+) at 15 K. For the FAD semiquinone, significantly different potentials were obtained by monitoring the titration at 100 or 293 K. Based on spectra at 293 K the N338T mutation shifted the first and second midpoint potentials for the FAD from +47 and -30 mV for wild type to -11 and -19 mV, respectively. The N338A mutation decreased the potentials to -37 and -49 mV. Lowering the midpoint potentials resulted in a decrease in the quinone reductase activity and negligible impact on disproportionation of ETF 1e (-) catalyzed by ETF-QO. These observations indicate that the FAD is involved in electron transfer to ubiquinone but not in electron transfer from ETF to ETF-QO. Therefore, the iron-sulfur cluster is the immediate acceptor from ETF.
 ...
PMID:The iron-sulfur cluster of electron transfer flavoprotein-ubiquinone oxidoreductase is the electron acceptor for electron transfer flavoprotein. 1867 1