In situ conversion of coproporphyrinogen to heme by murine mitochondria: terminal steps of the heme biosynthetic pathway.
Coproporphyrinogen oxidase (EC 1.3.3.3), protoporphyrinogen oxidase (EC 1.3.3.4), and ferrochelatase (EC 4.99.1.1) catalyze the terminal three steps of the heme biosynthetic pathway. All three are either bound to or associated with the inner mitochondrial membrane in higher eukaryotic cells. A current model proposes that these three enzymes may participate in some form of multienzyme complex with attendant substrate channeling (Grand-champ, B., Phung, N., & Nordmann, Y., 1978, Biochem. J. 176, 97-102; Ferreira, G.C., et al., 1988, J. Biol. Chem. 263, 3835-3839). In the present study we have examined this question in isolated mouse mitochondria using two experimental approaches: one that samples substrate and product levels during a timed incubation, and a second that follows dilution of radiolabeled substrate by pathway intermediates. When isolated mouse mitochondria are incubated with coproporphyrinogen alone there is an accumulation of free protoporphyrin. When Zn is added as a substrate for the terminal enzyme, ferrochelatase, along with coproporphyrinogen, there is formation of Zn protoporphyrin with little accumulation of free protoporphyrin. When EDTA is added to this incubation mixture with Zn, Zn protoporphyrin formation is eliminated and protoporphyrin is formed. We have examined the fate of radiolabeled substrates in vitro to determine if exogenously supplied pathway intermediates can compete with the endogenously produced compounds. The data demonstrate that while coproporphyrinogen is efficiently converted to heme in vitro when the pathway is operating below maximal capacity, exogenous protoporphyrinogen can compete with endogenously formed protoporphyrinogen in heme production.(ABSTRACT TRUNCATED AT 250 WORDS)
Proulx
K L
KL
Department of Microbiology, University of Georgia, Athens 30602-2605.
Woodard
S I
SI
Dailey
H A
HA
D000818:Animals; D003304:Coproporphyrinogen Oxidase / Q000378:metabolism; D003305:Coproporphyrinogens / Q000378:metabolism*; D005294:Ferrochelatase / Q000378:metabolism; D005420:Flavoproteins; D006418:Heme / Q000096:biosynthesis*; D008297:Male; D051379:Mice; D008811:Mice, Inbred DBA; D008813:Mice, Inbred ICR; D008930:Mitochondria, Liver / Q000378:metabolism*; D024101:Mitochondrial Proteins; D010088:Oxidoreductases / Q000378:metabolism; D044925:Oxidoreductases Acting on CH-CH Group Donors; D050768:Protoporphyrinogen Oxidase; D011524:Protoporphyrins / Q000096:biosynthesis / Q000378:metabolism; D015032:Zinc / Q000378:metabolism
D003305:Coproporphyrinogens; D005420:Flavoproteins; D024101:Mitochondrial Proteins; D011524:Protoporphyrins; C017803:zinc protoporphyrin; D006418:Heme; C011539:protoporphyrinogen; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; D003304:Coproporphyrinogen Oxidase; C493979:Ppox protein, mouse; D050768:Protoporphyrinogen Oxidase; D005294:Ferrochelatase; D015032:Zinc