Cloned (Comment) | Organism |
---|---|
hemeN gene, overexpression in Escherichia coli BL21(DE3) | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
C62S | inactive mutant, no Fe-S cluster formation | Escherichia coli |
C66S | inactive mutant, no Fe-S cluster formation | Escherichia coli |
C69S | inactive mutant, no Fe-S cluster formation | Escherichia coli |
C71S | inactive mutant, same Fe-S cluster formation as in wild-type HemN | Escherichia coli |
F68L | mutant with 89% of wild-type activity | Escherichia coli |
G111V/G113V | double mutation of Gly-111 and Gly-113, which are part of the potential GGGTP S-adenosyl-L-methionine binding motif, completely abolishes enzyme activity, reduced Fe-S cluster formation | Escherichia coli |
H58L | inactive mutant, no Fe-S cluster formation | Escherichia coli |
Y56F | mutant with 45% of wild-type activity and reduced Fe-S cluster formation | Escherichia coli |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
EDTA | strong inhibition | Escherichia coli | |
o-phenanthroline | strong inhibition | Escherichia coli |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
52000 | - |
gel filtration in combination with glycerol gradient centrifugation | Escherichia coli |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant HemN | Escherichia coli |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
coproporphyrinogen III + 2 S-adenosyl-L-methionine = protoporphyrinogen IX + 2 CO2 + 2 L-methionine + 2 5'-deoxyadenosine | catalytic, radical mechanism | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
coproporphyrinogen-III + S-adenosyl-L-methionine | HemN catalyzes the oxygen-independent conversion of coproporphyrinogen-III to protoporphyrinogen IX, requires S-adenosyl-L-methionine, NAD(P)H and additional cytoplasmatic components for catalysis. Cys-62, Cys-66 and Cys-69 are part of the conserved CXXXCXXC motif and essential for iron-sulfur cluster formation and enzyme function. Gly-111 and Gly-113 are part of the potential GGGTP S-adenosyl-L-methionine binding motif and essential for enzymatic function, catalytic, radical mechanism | Escherichia coli | protoporphyrinogen IX + CO2 + L-methionine + 5'-deoxyadenosine | - |
? |
Subunits | Comment | Organism |
---|---|---|
monomer | 1* 52734, sequence calculation | Escherichia coli |
Synonyms | Comment | Organism |
---|---|---|
More | Radical SAM enzyme | Escherichia coli |
oxygen-independent CPO | - |
Escherichia coli |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Escherichia coli |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
4Fe-4S-center | requirement, oxygen-sensitive Fe-S cluster, Cys-62, Cys-66 and Cys-69 are part of the conserved CXXXCXXC motif and essential for Fe-S cluster formation and enzyme function, Tyr-56 and His-58 are important for the Fe-S cluster integrity, His-58 may provide the fourth ligand besides the three cysteine residues | Escherichia coli | |
NADH | requires NAD(P)H, higher activity than with NADPH as cofactor | Escherichia coli | |
NADPH | requires NAD(P)H, lower activity than with NADH as cofactor | Escherichia coli | |
S-adenosyl-L-methionine | uses S-adenosyl-L-methionine as a cofactor | Escherichia coli |