Literature summary for 1.3.3.3 extracted from

Kim, D.H.; Hino, R.; Adachi, Y.; Kobori, A.; Taketani, S.
The enzyme engineering of mutant homodimer and heterodimer of coproporphyinogen oxidase contributes to new insight into hereditary coproporphyria and harderoporphyria (2013), J. Biochem., 154, 551-559 .

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant coexpression of HA-tagged wild-type enzyme from healthy donors and of His-tagged mutant enzyme from HCP patients in Escherichia coli strain BL21, the His-enzyme forms a heterodimer in association with the HA-tagged enzyme, The monomeric form of mutated CPOX does not show any activity and homodimeric enzymes derived from HCP mutant show low activity (about 20% of the control). The chimeric heterodimers with wild-type and mutated subunits from HCP patients show low protoporphyrinogen producing activity. The active site of the enzyme involved in the second step of decarboxylation is encoded in exon 6	Homo sapiens

Cloned (Comment)

Organism

recombinant coexpression of HA-tagged wild-type enzyme from healthy donors and of His-tagged mutant enzyme from HCP patients in Escherichia coli strain BL21, the His-enzyme forms a heterodimer in association with the HA-tagged enzyme, The monomeric form of mutated CPOX does not show any activity and homodimeric enzymes derived from HCP mutant show low activity (about 20% of the control). The chimeric heterodimers with wild-type and mutated subunits from HCP patients show low protoporphyrinogen producing activity. The active site of the enzyme involved in the second step of decarboxylation is encoded in exon 6

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
D400A	site-directed mutagenesis, the inactive mutant D400A remains in a monomeric form	Homo sapiens
G402D	site-directed mutagenesis, the mutant enzyme forms dimers	Homo sapiens
K404E	a naturally occuring mutant derived from patients with harderoporphyria, the mutant produces less harderoporphyrinogen. The K404E mutation leads to diminishment of the second step of the decarboxylation reaction during the conversion of coproporphyrinogen to protoporphyrinogen. The mutant enzyme forms dimers	Homo sapiens
K404H	site-directed mutagenesis, the mutant produces a high level of harderoporphyrinogen with low production of protoporphyrinogen similar to mutant K404E	Homo sapiens
K404Q	site-directed mutagenesis, the mutant shows unaltered activity compared to the wild-type enzyme	Homo sapiens
additional information	enzyme engineering of mutant homodimer and heterodimer of coproporphyinogen oxidase. The His-tagged mutant enzyme forms a heterodimer in association with the HA-tagged wild-type enzyme, The monomeric form of mutated CPOX does not show any activity and homodimeric enzymes derived from hereditary coproporphyria (HCP) mutant show low activity (about 20% of the control). The chimeric heterodimers with wild-type and mutated subunits from HCP patients show low protoporphyrinogen producing activity. Some mutations of amino acids 401-404 are associated with marked accumulation of reaction intermediate harderoporphyrinogen, with a decrease in the production of protoporphyrinogen, whereas K404E derived from patients with harderoporphyria produces less harderoporphyrinogen. Functional analysis of heterodimer of mutant/wild-type complex, heterophilic forms of His-R388W/HA-wild-type, His-R391W/HA-wild-type, His-D400A/HA-wild-type, His-R401W/HA-wild-type, His-G402D/HA-wild-type and His-K404E/HA-wild-type, overview	Homo sapiens
R388W	site-directed mutagenesis, the mutant enzyme forms dimers	Homo sapiens
R391W	site-directed mutagenesis, the mutant enzyme forms dimers	Homo sapiens
R401W	site-directed mutagenesis, the mutant enzyme forms dimers	Homo sapiens

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
74000	-	recombinant wild-type enzyme	Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
coproporphyrinogen III + O2 + 2 H+	Homo sapiens	via harderoporphyrinogen intermediate production, overall reaction	protoporphyrinogen-IX + 2 CO2 + 2 H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	P36551	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant HA-tagged wild-type enzyme and His-tagged mutant enzyme from Escherichia coli by nickel and heparin affinity chromatography, respectively	Homo sapiens

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
coproporphyrinogen III + O2 + 2 H+	via harderoporphyrinogen intermediate production, overall reaction	Homo sapiens	protoporphyrinogen-IX + 2 CO2 + 2 H2O	-	?

Subunits

Subunits	Comment	Organism
homodimer	2 * 37000, about, recombinant wild-type enzyme, SDS-PAGE	Homo sapiens

Synonyms

Synonyms	Comment	Organism
coproporphyinogen oxidase	-	Homo sapiens
CPOX	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.4	-	assay at	Homo sapiens

General Information

General Information	Comment	Organism
malfunction	partial deficiency of the enzyme coproporphyrinogen oxidase (CPOX) causes hereditary coproporphyria (HCP), an autosomal dominant-inherited disease of heme biosynthesis. Patients suffering HCP show 50% of normal activity and those with the rare autosomal recessive harderoporphyria accumulate harderoporphyrinogen, an intermediate porphyrin of the CPOX reaction. As only patients with mutation K404E in this region develop harderoporphyria, the K404E mutation leads to diminishment of the second step of the decarboxylation reaction during the conversion of coproporphyrinogen to protoporphyrinogen, implying that the active site of the enzyme involved in the second step of decarboxylation is encoded in exon 6	Homo sapiens
additional information	the active site of the enzyme involved in the second step of decarboxylation is encoded in exon 6. Aspartic acid at position 400 can be a prerequisite for dimerization of enzyme CPOX	Homo sapiens