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Applied and Environmental Microbiology, October 2002, p. 4894-4899, Vol. 68, No. 10
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.10.4894-4899.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Inactivation of the Elongation Factor Tu by Mosquitocidal Toxin-Catalyzed Mono-ADP-Ribosylation

Jörg Schirmer,1 Hans-Joachim Wieden,2 Marina V. Rodnina,2 and Klaus Aktories1*

Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, Albert-Ludwigs-Universität Freiburg, D-79104 Freiburg,1 Institute of Physical Biochemistry, University of Witten/Herdecke, D-58448 Witten, Germany2

Received 29 March 2002/ Accepted 17 July 2002

The mosquitocidal toxin (MTX) produced by Bacillus sphaericus strain SSII-1 is an ~97-kDa single-chain toxin which contains a 27-kDa enzyme domain harboring ADP-ribosyltransferase activity and a 70-kDa putative binding domain. Due to cytotoxicity toward bacterial cells, the 27-kDa enzyme fragment cannot be produced in Escherichia coli expression systems. However, a nontoxic 32-kDa N-terminal truncation of MTX can be expressed in E. coli and subsequently cleaved to an active 27-kDa enzyme fragment. In vitro the 27-kDa enzyme fragment of MTX ADP-ribosylated numerous proteins in E. coli lysates, with dominant labeling of an ~45-kDa protein. Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry combined with peptide mapping identified this protein as the E. coli elongation factor Tu (EF-Tu). ADP ribosylation of purified EF-Tu prevented the formation of the stable ternary EF-Tuaminoacyl-tRNAGTP complex, whereas the binding of GTP to EF-Tu was not altered. The inactivation of EF-Tu by MTX-mediated ADP-ribosylation and the resulting inhibition of bacterial protein synthesis are likely to play important roles in the cytotoxicity of the 27-kDa enzyme fragment of MTX toward E. coli.

* Corresponding author. Mailing address: Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, Albertstr. 25, D-79104 Freiburg, Germany. Phone: 49-761-2035301. Fax: 49-761-2035311. E-mail: aktories{at}uni-freiburg.de.

Applied and Environmental Microbiology, October 2002, p. 4894-4899, Vol. 68, No. 10
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.10.4894-4899.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.





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