Electron Transfer Flavoprotein Dehydrogenase Research Articles

The purification and some properties of electron transfer flavoprotein and general fatty acyl coenzyme A dehydrogenase from pig liver mitochondria.

Electron-transferring flavoprotein (ETF) and acyl dehydrogenases of pig liver mitochondria have been isolated in good yield by a new procedure. ETF and general acyl dehydrogenase appear homogenous, are free of reciprocal contamination, react with neither pyridine nucleotides not cytochrome c, and are completely dependent upon each other for reduction of dichlorophenol indophenol by acyl-CaA substrates. The properties of the present preparation (some of which differ significantly from those previously described) are presented. Sedimentation of ETF in 0.02 M KP-i yields a M-r for the native ETF of 58,00 plus or minus 3,000, whereas sedimentation of reduced and alkylated ETF in guanidine HCl yields a M-r of 26,000. Electrophoresis on sodium dodecyl sulfate gels in the presence or absence of mercaptoethanol gives a M-r of about 27,000 and flavin analysis gives a minimum molecular weight of about the same figure. Thus, ETF appears to contain one flavin (at least 90% FAD, by chromatographic and fluorescence characteristics) per 26,000 M-r, and therefore may be composed of two subunits with one flavin each. Sodium dodecyl sulfate gel electrophoresis of general acyl dehydrogenase in the absence of mercaptoethanol gives a band corresponding to a M-r of 84,000; in the presence of mercaptoethanol a band corresponding to a M-r of 42,000 is found. The minimum molecular weight based on flavin content is 40,500. These data considered in conjunction with previous reports from other laboratories, suggest a structure of four subunits per mol with one flavin per subunit..

Evidence That apo-Reduced Nicotinamide Adenine Dinucleotide Dehydrogenase and apo-Electron-transferring Flavoprotein from Peptostreptococcus elsdenii Are Identical

Abstract Electron-transferring flavoprotein (ETF) and NADH dehydrogenase, containing a novel orange flavin, have been highly purified from the strict anaerobe Peptostreptococcus elsdenii. Both enzymes couple the oxidation of NADH to the reduction of dyes but only ETF couples the oxidation of NADH to the reduction of butyryl coenzyme A dehydrogenase (ETF activity). Disc gel electrophoresis in sodium dodecyl sulfate and urea, immunological studies, and amino acid analyses have provided strong evidence that the apoproteins of ETF and NADH dehydrogenase are very similar or identical. The two enzyme preparations differ structurally in the proportions of their flavin chromophores, FAD and two novel modified flavins, 6-hydroxy-7,8-dimethyl-10-(ribityl-5'-ADP)-isoalloxazine and 7-methyl-8-hydroxy-10-(ribityl-5'-ADP)-isoalloxazine (8-OH-FAD). Preparations with ETF activity contain FAD as the predominant prosthetic group and low amounts of the two modified flavins. NADH dehydrogenase preparations contain all three flavins, about 50% FAD and 50% modified flavins; there is a high proportion of 8-OH-FAD which has an intense absorption band at 475 nm and imparts an orange color to the protein. Differences in catalytic activity, absorption spectrum, electrophoretic mobility, and isoelectric point of ETF and NADH dehydrogenase are most likely due to different proportions of the three flavins.