Proof for Substitution of Endogenous Iron (II) in Lipoxygenase by Exogenous Cu

• Published in: Applied biochemistry and biotechnology Vol. 162; no. 4; pp. 1147 - 1152
• Main Authors: Cai, Yan, Xu, Hong, Xia, Yongmei, Su, Yafen, Fang, Yun
• Format: Journal Article
• Language: English
• Published: New York : Humana Press Inc 2010
• Subjects: copper; enzyme activity; iron; lipoxygenase; metal ions; soybeans
• ISSN: 0273-2289; 1559-0291
• DOI: 10.1007/s12010-009-8876-5

Soybean lipoxygenase (LOX) contains endogenous iron (II) at the active site, which is important for the enzyme activity. The activity of LOX can be accelerated by some exogenous metal ions including Cu²⁺. However, the mechanism of the activity improvement caused by exogenous metal ions remains unclear, not only for LOX but for most other metalloenzymes. Meanwhile, the possibility that exogenous metal ions can displace endogenous iron (II) is still in discussion for a lack of a direct and quantitative proof. In this paper, a quantitative proof of replacing iron (II) inside LOX by exogenous Cu²⁺ was provided, simply using UV-Vis spectrometry with two indicators p-carboxylantipyrylazo and 9-(4-carboxyphenyl)-2,3,7-trihydroxyl-6-fluorine. A 0.56 μM free iron (II) was observed in the bulk solution after incubating 9.45 μM Cu²⁺ with 16.10 μM LOX at 20°C for 5 min, which is in coincidence with the decrement of Cu²⁺ in the bulk solution (0.53 μM), implying that iron (II) was replaced by Cu²⁺.