Details

Autor(en) / Beteiligte

• Philpott, C.C.
• Haile, D.
• Rouault, T.A.
• Klausner, R.D.

Titel

Modification of a free Fe-S cluster cysteine residue in the active iron-responsive element-binding protein prevents RNA binding

Ist Teil von

• The Journal of biological chemistry, 1993-08, Vol.268 (24), p.17655-17658

Ort / Verlag

Bethesda, MD: Elsevier Inc

Erscheinungsjahr

1993

Quelle

Free E-Journal (出版社公開部分のみ）

Beschreibungen/Notizen

• The iron-responsive element-binding protein (IRE-BP) binds to specific RNA stem-loop structures called iron-responsive elements (IREs), which mediate the post-transcriptional regulation of a variety of genes involved in iron metabolism. The IRE-BP is cytosolic aconitase, and a [4Fe-4S] cubane cluster is required for aconitase activity but is associated with loss of IRE binding affinity. Chemical modification of the IRE-BP can abrogate RNA binding and the 3 cysteines predicted to coordinate the Fe-S cluster in the IRE-BP could be targets for modification. We report the expression of recombinant IRE-BP in which the three putative cluster cysteines (Cys-437, Cys-503, and Cys-506) have been mutated to serine residues. Replacement of any or all of these cysteine residues results in a complete loss of aconitase activity. While all of the mutants bind RNA, substitution of Cys-437 specifically renders the IRE-BP resistant to inactivation by low concentrations of N-ethylmaleimide or diamide. These results identify Cys-437 as the target of in vitro regulation of RNA binding in the IRE-BP and suggest that, in the RNA-binding form of the protein, Cys-437 is free and therefore available for modifications that inhibit RNA binding.