Rat hepatic uroporphyrinogen III co-synthase. Purification and evidence for a bound folate coenzyme participating in the biosynthesis of uroporphyrinogen III.
Rat hepatic uroporphyrinogen III co-synthase was isolated and purified 73-fold with a 13% yield by (NH4)2SO4 fractionation and sequential chromatography on DEAE-Sephacel, Sephadex G-100 (superfine grade) and folate-AH-Sepharose 4B. The purified co-synthase has an Mr of approx. 42 000, and is resolved into two bands, each possessing co-synthase activity, by polyacrylamide-gel electrophoresis. A factor was dissociated from the purified co-synthase. Results of both microbiological and competitive protein-binding assays suggest that it is a pteroylpolyglutamate. The isolated pteroylpolyglutamate factor was co-eluted with authentic N5-methyltetrahydropteroylheptaglutamate on DEAE-Sephacel. Uroporphyrinogen III is formed by cosynthase-free preparations of uroporphyrinogen I synthase in the presence of tetrahydropteroylglutamate. Tetrahydropeteroylheptaglutamate is also able to direct the formation of equivalent amounts of uroporphyrinogen III at a concentration approximately one-hundredth that of tetrahydropteroylmonoglutamate. These results suggest that a reduced pteroylpolyglutamate factor is associated with rat hepatic uroporphyrinogen III co-synthase, and that this may function as a coenzyme for the biosynthesis of uroporphyrinogen III.
D000818:Animals; D002845:Chromatography; D003067:Coenzymes / Q000378:metabolism*; D003600:Cytosol / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D005492:Folic Acid / Q000378:metabolism*; D011163:Hydroxymethylbilane Synthase / Q000378:metabolism; D007535:Isomerases / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight; D011624:Pteroylpolyglutamic Acids / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D014576:Uroporphyrinogen III Synthetase / Q000302:isolation & purification / Q000378:metabolism*
D003067:Coenzymes; D011624:Pteroylpolyglutamic Acids; D005492:Folic Acid; D011163:Hydroxymethylbilane Synthase; D014576:Uroporphyrinogen III Synthetase; D007535:Isomerases