Molecular cloning and tissue distribution of rat sarcosine dehydrogenase.
Sarcosine dehydrogenase (SarDH) is a mitochondrial flavoenzyme involved in the oxidative degradation of choline to glycine. The absence of SarDH activity in humans is genetically transmitted and is the cause of an amino acid metabolism disorder called sarcosinemia. Tryptic fragments of the purified enzyme from rat liver were subjected to Edman degradation and the sequences obtained were used to clone the cDNA encoding the full length protein. The deduced amino acid sequence of SarDH shares an overall similarity of 47% with dimethylglycine dehydrogenase (Me2GlyDH), another flavoenzyme involved in the mitochondrial choline catabolism with a similar FAD-binding domain. Covalent binding of FAD to SarDH was demonstrated by the observation of strong fluorescence at 530 nm under excitation at 450 nm of the enzyme immunoprecipitated under denaturing conditions from liver extracts. The localization of SarDH immunoreactivity in the mitochondrial matrix was confirmed by Western-blot analysis of purified mitochondrial fractions. Finally, the tissue distribution of SarDH was investigated by Northern-blot analysis of total RNA and Western-blot analysis of total protein from several rat tissues. A strong expression in the liver, but also in the lung, pancreas, kidney, thymus, and oviduct was observed. We therefore suggest that the enzymes of the choline catabolism pathway are important also for metabolism in nonhepatic tissues.
Bergeron
F
F
INSERM U376, CHU Arnaud de Villeneuve, Montpellier, France.
Otto
A
A
Blache
P
P
Day
R
R
Denoroy
L
L
Brandsch
R
R
Bataille
D
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary; D006801:Humans; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology; D008969:Molecular Sequence Data; D010089:Oxidoreductases, N-Demethylating / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D050892:Sarcosine Dehydrogenase; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D010089:Oxidoreductases, N-Demethylating; C494395:SARDH protein, human; D050892:Sarcosine Dehydrogenase; C494397:Sardh protein, rat