Hammarberg T, Zhang Y Y, Lind B, Radmark O, Samuelsson B
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Eur J Biochem. 1995 Jun 1;230(2):401-7. doi: 10.1111/j.1432-1033.1995.0401h.x.
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.
对人5-脂氧合酶中的非血红素铁中心进行了研究。重组酶在大肠杆菌中表达，经纯化后测定铁含量和酶活性。对于未突变的酶，铁含量为1.01±0.19摩尔/摩尔。C末端Ile673的缺失导致铁含量为0.03±0.07摩尔/摩尔，且脂氧合酶活性无法检测到。His367、Glu376和Asn554的突变导致酶活性大幅降低（<未突变对照的2%），但铁仍然存在。除了Glu376，5-脂氧合酶中的其他八个保守酸性残基（Asp/Glu）被替换，其中没有一个对酶活性至关重要。我们得出结论，Ile673是5-脂氧合酶中的铁配体，而我们的结果不支持Glu376或Asn554具有此功能。讨论了His367作为可替代铁配体的可能作用。
