title: Mutations at the C-terminal isoleucine and other potential iron ligands of 5-lipoxygenase.
abstract: The non-heme iron centre in human 5-lipoxygenase was studied. Recombinant enzyme was expressed in Escherichia coli, purified and assayed for iron content and enzyme activity. For non-mutated enzyme, the iron content was 1.01 +/- 0.19 mol/mol. Deletion of the C-terminal Ile673 resulted in an iron content of 0.03 +/- 0.07 mol/mol and undetectable lipoxygenase activity. Mutations at His367, Glu376 and Asn554 led to drastically decreased enzyme activity (< 2% of non-mutated control) but iron was still present. In addition to Glu376, eight other conserved acidic residues (Asp/Glu) in 5-lipoxygenase were replaced, none of which was crucial for enzyme activity. We conclude that Ile673 is an iron ligand in 5-lipoxygenase, while our results do not support that Glu376 or Asn554 have this function. The possible role of His367 as a replaceable iron ligand is discussed.
authors.affiliation: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
authors.forename: T
authors.initials: T
authors.lastname: Hammarberg
authors.forename: Y Y
authors.initials: YY
authors.lastname: Zhang
authors.forename: B
authors.initials: B
authors.lastname: Lind
authors.forename: O
authors.initials: O
authors.lastname: Radmark
authors.forename: B
authors.initials: B
authors.lastname: Samuelsson
mesh_terms: D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007501:Iron / Q000378:metabolism*; D007532:Isoleucine / Q000235:genetics* / Q000378:metabolism; D008024:Ligands; D016297:Mutagenesis, Site-Directed
chemical_list: D008024:Ligands; D007532:Isoleucine; D007501:Iron; D001094:Arachidonate 5-Lipoxygenase