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Literature summary extracted from

  • Starbird, C.A.; Maklashina, E.; Sharma, P.; Qualls-Histed, S.; Cecchini, G.; Iverson, T.M.
    Structural and biochemical analyses reveal insights into covalent flavinylation of the Escherichia coli Complex II homolog quinol fumarate reductase (2017), J. Biol. Chem., 292, 12921-12933 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.3.5.1 operon FrdABCD, recombinant expression of wild-type and mutant FrdAs in Escherichia coli strain DW35, in which both the frd and sdh operons are disrupted via the insertion of a kanamycin gene Escherichia coli

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.3.5.1 purified FrdA mutant E245Q, hanging-drop vapor diffusion method, mixing of 0.001 ml of 15 mg/ml protein in 25 mM Tris-HCl, pH 7.4, 1 mM EDTA, 0.02% C12E9, with 0.001 ml of reservoir solution containing 275mM sodium malonate, 19% PEG 6000, 100 mM sodium citrate, pH 4.0, 1 mM EDTA, and 0.001% dithiothreitol, and equilibration against 1 ml of reservoir solution, 20°C, X-ray diffraction structure determination and analysis at 4.25 A resolution Escherichia coli

Protein Variants

EC Number Protein Variants Comment Organism
1.3.5.1 D288N site-directed mutagenesis, the FrdAD288N variant shows minimal residual fluorescence, suggesting covalent flavinylation is severely compromised Escherichia coli
1.3.5.1 E245Q site-directed mutagenesis, the mutant of FrdA shows almost complete absence of covalent flavinylation. Mutant crystal structure analysis, overview Escherichia coli
1.3.5.1 H355S site-directed mutagenesis, the mutant of FrdA shows almost complete absence of covalent flavinylation Escherichia coli
1.3.5.1 additional information circular dichroism spectroscopy of wild-type and variant FrdA subunits, measurement of flavin in wild-type and variant FrdA subunits and determination of the quantity of flavin covalently associated with FrdA, overview Escherichia coli
1.3.5.1 R287K site-directed mutagenesis, the mutant of FrdA shows almost complete absence of covalent flavinylation Escherichia coli
1.3.5.1 R390K site-directed mutagenesis, the mutant of FrdA shows almost complete absence of covalent flavinylation Escherichia coli
1.3.5.1 R390Q site-directed mutagenesis, the mutant of FrdA shows almost complete absence of covalent flavinylation Escherichia coli

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.3.5.1 malonate
-
Escherichia coli
1.3.5.1 oxaloacetate
-
Escherichia coli

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.3.5.1 succinate + menaquinone Escherichia coli
-
fumarate + menaquinol
-
?

Organism

EC Number Organism UniProt Comment Textmining
1.3.5.1 Escherichia coli P00363
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.3.5.1 soluble recombinant enzyme mutant E245Q from Escherichia coli strain DW35 by anion exchange chromatography, ultrafitration, a second different step of anion exchange chromatography, followed by gel filtration Escherichia coli

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.3.5.1 succinate + menaquinone
-
Escherichia coli fumarate + menaquinol
-
?
1.3.5.1 succinate + menaquinone QFR can also catalyze the reverse reaction, succinate oxidation, albeit with slower kinetics and poorer catalytic efficiency Escherichia coli fumarate + menaquinol
-
r

Synonyms

EC Number Synonyms Comment Organism
1.3.5.1 Complex II homolog
-
Escherichia coli
1.3.5.1 FRdABCD
-
Escherichia coli
1.3.5.1 menaquinol-1 fumarate reductase
-
Escherichia coli
1.3.5.1 QFR
-
Escherichia coli
1.3.5.1 quinol:fumarate reductase
-
Escherichia coli

Cofactor

EC Number Cofactor Comment Organism Structure
1.3.5.1 FAD covalently attached FAD within the FrdA subunit, the SdhE assembly factor enhances covalent flavinylation of complex II homologues. Analysis of the mechanisms of covalent flavinylation of FrdA subunit, and propose of chemical mechanism of covalent flavinylation via a quinone-methide intermediate, overview. In Escherichia coli QFR, the flavin is covalently attached to the FrdA subunit via an 8x02 linkage to the N-epsilon of His44. This places the flavin isoalloxazine group at the interface of two domains, termed the flavin-binding domain and the capping domain. These domains can move with respect to each other, which may allow the catalytic subunit to close over the substrate-bound active site and protect the reaction intermediates from solvent. Quantitation of covalent flavinylation under aerobic and anaerobic growth conditions Escherichia coli
1.3.5.1 additional information the SdhE assembly factor binds directly to the FrdA subunit prior to assembly into the intact complex. The interaction involves a surface of SdhE containing a conserved RGXXE motif. SdhE is required for enzyme complex function, it promotes covalent flavinylation of FrdA subunit Escherichia coli

General Information

EC Number General Information Comment Organism
1.3.5.1 evolution enzyme QFR is a member of the complex II superfamily and is composed of FrdABCD subunits Escherichia coli
1.3.5.1 physiological function quinol:fumarate reductase (QFR, FrdABCD) catalyzes the interconversion of fumarate and succinate at a covalently attached FAD within the FrdA subunit. The SdhE assembly factor enhances covalent flavinylation of complex II homologues, mechanism, overview. QFR catalyzes the reduction of fumarate (kcat = 250/s) at this flavin-based active site during anaerobic respiration with fumarate as the terminal electron acceptor. In this process, the two electrons for fumarate reduction derived from the oxidation of menaquinol in the membrane and the two protons are likely transferred from solvent via a proton shuttle pathway consisting of the FrdA-E245, FrdA-R248, and FrdA-R287 side chains located on the capping domain. QFR can also catalyze the reverse reaction, succinate oxidation, albeit with slower kinetics (kcatx02= 30/s) and poorer catalytic efficiency Escherichia coli