Flavin adenine dinucleotide and flavin mononucleotide metabolism in rat liver--the occurrence of FAD pyrophosphatase and FMN phosphohydrolase in isolated mitochondria.
In order to gain some insight into mitochondrial flavin biochemistry, rat liver mitochondria essentially free of lysosomal and microsomal contamination were prepared and their capability to metabolise externally added and endogenous FAD and FMN tested both spectroscopically and via HPLC. The existence of two novel mitochondrial enzymes, namely FAD pyrophosphatase (EC 3.6.1.18) and FMN phosphohydrolase (EC 3.1.3.2), which catalyse FAD-->FMN and FMN-->riboflavin conversion, respectively, is shown. They differ from each other and from extramitochondrial enzymes, as judged by their pH profile and inhibitor sensitivity, and can be separated in a partial FAD pyrophosphatase purification. Digitonin titration and subfractionation experiments show that FAD pyrophosphatase is located in the outer mitochondrial membrane and FMN phosphohydrolase in the intermembrane space. Since these enzymes can metabolise endogenous FAD and FMN, which are made available by using both Triton X-100 and the effector oxaloacetate, a proposal is made that FAD pyrophosphatase and FMN phosphohydrolase play a major role in mitochondrial flavoprotein turnover.
Barile
M
M
Dipartimento di Biochimica e Biologia Molecolare, Università di Bari, and Centro di Studio sui Mitocondri e Metabolismo Energetico, C.N.R., Italy. barile@bioserver.uniba.it
Brizio
C
C
De Virgilio
C
C
Delfine
S
S
Quagliariello
E
E
Passarella
S
S
D000135:Acid Phosphatase / Q000302:isolation & purification / Q000378:metabolism*; D000249:Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D002458:Cell Fractionation; D002851:Chromatography, High Pressure Liquid; D005486:Flavin Mononucleotide / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism*; D005420:Flavoproteins / Q000302:isolation & purification / Q000378:metabolism; D005453:Fluorescence; D006868:Hydrolysis; D007700:Kinetics; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology* / Q000378:metabolism; D009708:Nucleotidases / Q000378:metabolism*; D010071:Oxaloacetates / Q000494:pharmacology; D011755:Pyrophosphatases / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D005420:Flavoproteins; D010071:Oxaloacetates; D005182:Flavin-Adenine Dinucleotide; D000249:Adenosine Monophosphate; D005486:Flavin Mononucleotide; C033670:FMN phosphatase; D009708:Nucleotidases; D000135:Acid Phosphatase; D011755:Pyrophosphatases; C023417:FAD pyrophosphatase