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Plant Physiol, February 2001, Vol. 125, pp. 943-954

Purification and Characterization of the Preprotein Translocase of the Outer Mitochondrial Membrane from Arabidopsis. Identification of Multiple Forms of TOM201

Wolf Werhahn, Astrid Niemeyer, Lothar Jänsch,2 Volker Kruft, Udo K. Schmitz, and Hans-Peter Braun*

Institut für Angewandte Genetik, Universität Hannover, Herrenhäuser Strasse 2, D-30419 Hannover, Germany (W.W., A.N., L.J., U.K.S., H.-P.B.); and Applied Biosystems, Paul-Ehrlich Strasse 17, 63225 Langen, Germany (V.K.)

The translocase of the outer mitochondrial membrane (TOM) complex is a preprotein translocase that mediates transport of nuclear-encoded mitochondrial proteins across the outer mitochondrial membrane. Here we report the purification of this protein complex from Arabidopsis. On blue-native gels the Arabidopsis TOM complex runs at 230 kD and can be dissected into subunits of 34, 23, 21, 8, 7, and 6 kD. The identity of four subunits could be determined by immunoblotting and/or direct protein sequencing. The 21- and the 23-kD subunits exhibit significant sequence homology to the TOM20 preprotein receptor from other organisms. Analysis by two-dimensional isoelectric focusing/Tricine sodium dodecyl sulfide-polyacrylamide gel electrophoresis revealed the presence of further forms for Arabidopsis TOM20. All TOM20 proteins comprise a large cytoplasmically exposed hydrophilic domain, which is degraded upon trypsination of intact mitochondria. Clones encoding four different forms of Arabidopsis TOM20 were identified and sequenced. The deduced amino acid sequences are rather conserved in the N-terminal half and in the very C-terminal part, but include a highly variable glycine-rich region close to the C terminus. Implications on the function of plant TOM complexes are discussed. Based on peptide and nucleic acid sequence data, the primary structure for Arabidopsis TOM40 is presented.

1 This work was supported by the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie.

2 Present address: Gesellschaft für Biotechnologische Forschung, Mascheroder Weg 1, 38124 Braunschweig, Germany.

* Corresponding author; e-mail Hans-Peter.Braun{at}mbox.genetik.uni-hannover.de; fax 49511-7623608.

© 2001 American Society of Plant Physiologists


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