The effects of pH, ionic strength, and chemical modifications on the reaction of electron transfer flavoprotein with an acyl coenzyme A dehydrogenase.
The effects of pH and ionic strength on the steady state kinetic parameters for reduction of electron transfer flavoprotein (ETF) by general acyl-CoA dehydrogenase were determined. The effect of pH on the turnover number (TN) of the reaction indicates the participation of an essential base with a pK alpha of 6.9. The KmETF of the dehydrogenase is invariant between pH 5.4 and 8.5, but increases 40-fold between pH 8.5 and 9.8. The parameter TN/KmETF follows the limiting Bronsted equation (In TN/KmETF = ln ko + 2.34ZAZB I 1/2) at ionic strength values between 0.01 and 0.125 M, indicating complementary charge interactions between the two flavoproteins. Covalent modifications of amino groups of ETF with trinitrobenzene sulfonate and acetic anhydride remove positive charges and result in an increase in KmETF of the dehydrogenase with no change of TN. However, exhaustive acetimidation of ETF amino groups, which maintains cationic charge at modified loci, does not alter the steady state kinetic parameters of the reaction. These results, in conjunction with previous chemical covalent modifications of dehydrogenase carboxyl residues (Frerman, F. E., Mielke, D., and Huhta, K. (1980) J. Biol. Chem. 255, 2199-2202), indicate that general acyl-CoA dehydrogenase and ETF interact in an electrostatic manner.
D044944:Acyl-CoA Dehydrogenases / Q000378:metabolism*; D000818:Animals; D010856:Columbidae; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D044927:Electron-Transferring Flavoproteins; D006863:Hydrogen-Ion Concentration; D007460:Iodoacetamide / Q000494:pharmacology; D007506:Iron-Sulfur Proteins; D007700:Kinetics; D008930:Mitochondria, Liver / Q000201:enzymology*; D009097:Multienzyme Complexes; D009994:Osmolar Concentration; D010088:Oxidoreductases / Q000378:metabolism*; D000587:Oxidoreductases Acting on CH-NH Group Donors; D014302:Trinitrobenzenesulfonic Acid / Q000494:pharmacology
D044927:Electron-Transferring Flavoproteins; D007506:Iron-Sulfur Proteins; D009097:Multienzyme Complexes; D014302:Trinitrobenzenesulfonic Acid; D004228:Dithionitrobenzoic Acid; D010088:Oxidoreductases; D044944:Acyl-CoA Dehydrogenases; D000587:Oxidoreductases Acting on CH-NH Group Donors; C023120:electron-transferring-flavoprotein dehydrogenase; D007460:Iodoacetamide