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Journal of Bacteriology, February 2003, p. 1174-1180, Vol. 185, No. 4
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.4.1174-1180.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Interactions between Phage-Shock Proteins in Escherichia coli

Hendrik Adams,{dagger} Wieke Teertstra, Jeroen Demmers, Rolf Boesten, and Jan Tommassen*

Department of Molecular Microbiology and Institute of Biomembranes, Utrecht University, 3584 CH Utrecht, The Netherlands

Received 23 July 2002/ Accepted 18 November 2002

Expression of the pspABCDE operon of Escherichia coli is induced upon infection by filamentous phage and by many other stress conditions, including defects in protein export. Expression of the operon requires the alternative sigma factor {sigma}54 and the transcriptional activator PspF. In addition, PspA plays a negative regulatory role, and the integral-membrane proteins PspB and PspC play a positive one. In this study, we investigated whether the suggested protein-protein interactions implicated in this complex regulatory network can indeed be demonstrated. Antisera were raised against PspB, PspC, and PspD, which revealed, in Western blotting experiments, that PspC forms stable sodium dodecyl sulfate-resistant dimers and that the hypothetical pspD gene is indeed expressed in vivo. Fractionation experiments showed that PspD localizes as a peripherally bound inner membrane protein. Cross-linking studies with intact cells revealed specific interactions of PspA with PspB and PspC, but not with PspD. Furthermore, affinity-chromatography suggested that PspB could bind PspA only in the presence of PspC. These data indicate that regulation of the psp operon is mediated via protein-protein interactions.

* Corresponding author. Mailing address: Department of Molecular Microbiology, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands. Phone: (31) 30 253 2999. Fax: (31) 30 251 3655. E-mail: J.P.M.Tommassen{at}bio.uu.nl.

{dagger} Present address: École Supérieure de Biotechnologie de Strasbourg, 67400 Illkirch, France.

Journal of Bacteriology, February 2003, p. 1174-1180, Vol. 185, No. 4
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.4.1174-1180.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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