The terminal two enzymes of the heme biosynthetic pathway, protoporphyrinogen oxidase and ferrochelatase, catalyze the conversion of protoporphyrinogen to protoporphyrin followed by insertion of ferrous iron to form the end product protoheme. These enzymes are of general biochemical interest for not only their unique catalytic functions, but also because of their vectoral organization across the membrane. Medically they are of interest since decreased activities of either one results in a disease condition known generally as porphyria. In addition it has been shown recently that drug induced heme destruction in cytochrome P450 can lead to a porphyric condition in healthy animals and that the mechanism of this porphyria involves in vivo inhibition ferrochelatase. The proposed research is designed to increase our understanding of ferrochelatase and protoporphyrinogen oxidase. Specifically the data gathered will include information about protein sequence, membrane topology of the proteins, details of protein-protein interactions and the mechanism of substrate and product transport into and out the mitochondrion. An additional benefit of the experimental approaches to be exploited in this study will be the design of ELISA assays to quantitate both ferrochelatase and protoporphyrinogen oxidase. These procedures should prove to be of general medical use in future screening for varigate porphyria and protoporphyria. The major experimental approaches can be broken down into four categories. These are: 1) examination of possible protein-protein interactions between ferrochelatase and protoporphyrinogen oxidase, 2) determination of the membrane topology of these two enzymes, 3) amino acid sequence determination of putative active site peptides for both enzymes and eventual total amino acid sequence for ferrochelatase and 4) examination of porphyrin and heme movement across the inner mitochondrial membrane.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK032303-04
Application #
3230730
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1983-04-01
Project End
1991-03-31
Budget Start
1986-04-01
Budget End
1987-03-31
Support Year
4
Fiscal Year
1986
Total Cost
Indirect Cost
Name
University of Georgia
Department
Type
Schools of Arts and Sciences
DUNS #
City
Athens
State
GA
Country
United States
Zip Code
30602
Dailey, Harry A; Meissner, Peter N (2013) Erythroid heme biosynthesis and its disorders. Cold Spring Harb Perspect Med 3:a011676
Medlock, Amy E; Najahi-Missaoui, Wided; Ross, Teresa A et al. (2012) Identification and characterization of solvent-filled channels in human ferrochelatase. Biochemistry 51:5422-33
Hamza, Iqbal; Dailey, Harry A (2012) One ring to rule them all: trafficking of heme and heme synthesis intermediates in the metazoans. Biochim Biophys Acta 1823:1617-32
Chen, Caiyong; Samuel, Tamika K; Sinclair, Jason et al. (2011) An intercellular heme-trafficking protein delivers maternal heme to the embryo during development in C. elegans. Cell 145:720-31
Dailey, Harry A; Septer, Alecia N; Daugherty, Lauren et al. (2011) The Escherichia coli protein YfeX functions as a porphyrinogen oxidase, not a heme dechelatase. MBio 2:e00248-11
Boynton, Tye O; Gerdes, Svetlana; Craven, Sarah H et al. (2011) Discovery of a gene involved in a third bacterial protoporphyrinogen oxidase activity through comparative genomic analysis and functional complementation. Appl Environ Microbiol 77:4795-801
Dailey, Tamara A; Boynton, Tye O; Albetel, Angela-Nadia et al. (2010) Discovery and Characterization of HemQ: an essential heme biosynthetic pathway component. J Biol Chem 285:25978-86
Chen, Wen; Dailey, Harry A; Paw, Barry H (2010) Ferrochelatase forms an oligomeric complex with mitoferrin-1 and Abcb10 for erythroid heme biosynthesis. Blood 116:628-30
Boynton, Tye O; Daugherty, Lauren E; Dailey, Tamara A et al. (2009) Identification of Escherichia coli HemG as a novel, menadione-dependent flavodoxin with protoporphyrinogen oxidase activity. Biochemistry 48:6705-11
Shepherd, M; Dailey, H A (2009) Peroxidase activity of cytochrome C facilitates the protoporphyrinogen oxidase reaction. Cell Mol Biol (Noisy-le-grand) 55:6-14

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