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Cloning and characterization of chironomidae ferrochelatase: Copper activation of the purified ferrochelatase

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Abstract

All organisms utilize ferrochelatase (EC 4.99.1.1) to catalyze the insertion of ferrous iron into protoposphyrin IX in the terminal step of the heme biosynthetic pathway. Different metal-binding affinity for the enzyme leads to changes in enzyme activity. In this work, we have cloned and over-experessed the enzyme from chironomidae in E. coli. The enzyme was purified and characterized. The recombinant enzyme showed higher enzymatic activity (four-fold increase) in the presence of copper ions and unaffected by calcium ions. Other divalent metal ions including magnesium, manganese, lead, reduced the enzyme activity by >60%. Over 90% of the enzyme activity was inhibited by Zn2+. The sequence alignment of amino acid residues reveals 83% homology with other ferrochelatases. The results of electron proton resonance (EPR) analysis showed that Fe2+ ion was present in the cluster of the recombinant enzyme complex. The recombinant enzyme also contained the [2Fe-2S] center with two-fold higher enzymatic activity than human ferrochelatase. (Mol Cell Biochem 262: 225–231, 2004)

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Authors and Affiliations

  1. Department of Biochemistry, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong

    Yuet Kin Leung, Kwong Fai Wong & John W. Ho

  2. Department of Chemistry, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong

    Hung Kay Lee

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  1. Yuet Kin Leung
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  2. Kwong Fai Wong
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Leung, Y.K., Wong, K.F., Lee, H.K. et al. Cloning and characterization of chironomidae ferrochelatase: Copper activation of the purified ferrochelatase. Mol Cell Biochem 262, 225–231 (2004). https://doi.org/10.1023/B:MCBI.0000038238.27488.9f

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