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Published April 9, 2004 | Published
Journal Article Open

A Monomer-to-Trimer Transition of the Human Mitochondrial Transcription Termination Factor (mTERF) Is Associated with a Loss of in Vitro Activity

Abstract

The human mitochondrial transcription termination factor (mTERF) is a nuclear-encoded 39-kDa protein that recognizes a mtDNA segment within the mitochondrial tRNALeu(UUR) gene immediately adjacent to and downstream of the 16 S rRNA gene. Binding of mTERF to this site promotes termination of rDNA transcription. Despite the fact that mTERF binds DNA as a monomer, the presence in its sequence of three leucine-zipper motifs suggested the possibility of mTERF establishing intermolecular interactions with proteins of the same or different type. When a mitochondrial lysate from HeLa cells was submitted to gel filtration chromatography, mTERF was eluted in two peaks, as detected by immunoblotting. The first peak, which varied in proportion between 30 and 50%, appeared at the position expected from the molecular mass of the monomer (41 ± 2 kDa), and the gel filtration fractions that contained it exhibited DNA binding activity. Most interestingly, the material in this peak had a strong stimulating activity on in vitro transcription of the mitochondrial rDNA. The second peak eluted at a position corresponding to an estimated molecular mass of 111 ± 5 kDa. No mTERF DNA binding activity could be detected in the corresponding gel filtration fractions. Therefore, we propose that mTERF exists in mitochondria in two forms, an active monomer and an inactive large size complex. The estimated molecular weight of this complex and the fact that purified mTERF can be eluted from a gel filtration column as a complex of the same molecular weight strongly suggest that this inactive complex is a homotrimer of mTERF.

Additional Information

© 2004 The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, November 17, 2003, and in revised form, January 25, 2004. We thank Arger Drew, Maria del Mar Roldan, and Jennifer Fish for expert technical assistance. We are very grateful to Petr Héyek, Patricio Fernandez-Silva, Miguel Martin-Hernandez, and Michael P. Murphy for valuable discussions. This work was supported by National Institutes of Health Grant GM11726 (to G. A.) 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.

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