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J. Biol. Chem., Vol. 278, Issue 25, 22325-22330, June 20, 2003

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Redox Regulation of 3'-Phosphoadenylylsulfate Reductase from Escherichia coli by Glutathione and Glutaredoxins^*

Christopher Horst Lillig  , Aristi Potamitou , Jens-Dirk Schwenn , Alexios Vlamis-Gardikas  and Arne Holmgren  ¶

From the Medical Nobel Institute for Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Scheeles Väg 2, 17177 Stockholm, Sweden and Biochemistry of Plants, Faculty for Biology, Ruhr-University Bochum, Universitätsstrasse 150, 44780 Bochum, Germany

Inorganic sulfate (, S^+VI) is reduced in vivo to sulfite (, S^+IV) via phosphoadenylylsulfate (PAPS) reductase. Escherichia coli lacking glutathione reductase and glutaredoxins (gor^–grxA^–grxB^–grxC^–) barely grows on sulfate. We found that incubation of PAPS reductase with oxidized glutathione leads to enzyme inactivation with simultaneous formation of a mixed disulfide between glutathione and the active site Cys-239. A newly developed method based on thiol-specific fluorescent alkylation and gel electrophoresis showed that glutathionylated PAPS reductase is reduced by glutaredoxins via a monothiol mechanism. This glutathionylated species was also observed in poorly growing gor^–grxA^–grxB^–grxC^– cells expressing inactive glutaredoxin 2 (Grx2) C9S/C12S. However, it was absent in better growing cells expressing monothiol Grx2 C12S or wild type Grx2. Reversible glutathionylation may thus regulate the activity of PAPS reductase in vivo.

Received for publication, March 5, 2003 , and in revised form, April 2, 2003.

^* This investigation was supported by grants from the Deutsche Forschungsgemeinschaft, the Karolinska Institute, the Knut and Alice Wallenberg Foundation, the Swedish Cancer Society (961), and the Wenner-Gren foundation. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ To whom correspondence should be addressed. Tel.: 46-8-728-7728; Fax: 46-8-728-4716; E-mail: arne.holmgren{at}mbb.ki.se.

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