Properties of the Cysteine Residues and the Iron-Sulfur Cluster of the Assimilatory 5'-Adenylyl Sulfate Reductase from Enteromorpha intestinalis
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- 作者：Sung-Kun Kim ; Afroza Rahman ; Richard C. Conover ; Michael K. Johnson ; Jeremy T. Mason ; Varinnia Gomes ; Masakazu Hirasawa ; Mace L. Moore ; Thomas Leustek ; and David B. Knaff
- 刊名：Biochemistry
- 出版年：2006
- 出版时间：April 18, 2006
- 年：2006
- 卷：45
- 期：15
- 页码：5010 - 5018
- 全文大小：464K
- 年卷期：v.45,no.15(April 18, 2006)
- ISSN：1520-4995
文摘
The 5'-adenylyl sulfate (APS) reductase from the marine macrophytic green alga Enteromorphaintestinalis uses reduced glutathione as the electron donor for the reduction of APS to 5'-AMP and sulfite.The E. intestinalis enzyme (EiAPR) is composed of a reductase domain and a glutaredoxin-like C-terminaldomain. The enzyme contains a single [4Fe-4S] cluster as its sole prosthetic group. Three of the enzyme'seight cysteine residues (Cys166, Cys257, and Cys260) serve as ligands to the iron-sulfur cluster. Site-directed mutagenesis experiments and resonance Raman spectroscopy are consistent with the presence ofa cluster in which only three of the four ligands to the cluster irons contributed by the protein are cysteineresidues. Site-directed mutagenesis experiments suggest that the thiol group of Cys250, a residue foundonly in algal APS reductases, is not an absolute requirement for activity. The other four cysteines that donot serve as cluster ligands, all of which are required for activity, are involved in the formation of tworedox-active disulfide/dithiol couples. The couple involving Cys342 and Cys345 has an Em value at pH7.0 of -140 mV, and the one involving Cys165 and Cys285 has an Em value at pH 7.0 of -290 mV. TheC-terminal portion of EiAPR, expressed separately, exhibits the cystine reductase activity characteristicof glutaredoxins. It is proposed that the Cys342-Cys345 disulfide provides the site for entry of electronsfrom reduced glutathione and that the Cys166-Cys285 disulfide may serve as a structural element thatis essential for keeping the enzyme in the catalytically active conformation.