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..
D015086:2,6-Dichloroindophenol; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D003065:Coenzyme A; D004579:Electron Transport; D044943:Fatty Acid Desaturases / Q000302:isolation & purification / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism; D005420:Flavoproteins / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D013053:Spectrophotometry; D013552:Swine
D000596:Amino Acids; D005227:Fatty Acids; D005420:Flavoproteins; D015086:2,6-Dichloroindophenol; D044943:Fatty Acid Desaturases; D003065:Coenzyme A