A monomer-to-trimer transition of the human mitochondrial transcription termination factor (mTERF) is associated with a loss of in vitro activity.
The human mitochondrial transcription termination factor (mTERF) is a nuclear-encoded 39-kDa protein that recognizes a mtDNA segment within the mitochondrial tRNA(Leu(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.
Asin-Cayuela
Jordi
J
Division of Biology, California Institute of Technology, Pasadena, California 91125, USA. attardi@caltech.edu
Helm
Mark
M
Attardi
Giuseppe
G
D020816:Amino Acid Motifs; D050976:Basic-Leucine Zipper Transcription Factors; D002850:Chromatography, Gel; D004247:DNA / Q000737:chemistry; D004272:DNA, Mitochondrial / Q000737:chemistry; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D016350:Leucine Zippers; D024101:Mitochondrial Proteins; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012336:RNA, Ribosomal, 16S / Q000737:chemistry; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism; D014158:Transcription, Genetic
D050976:Basic-Leucine Zipper Transcription Factors; D004272:DNA, Mitochondrial; C105178:MTERF1 protein, human; D024101:Mitochondrial Proteins; D012336:RNA, Ribosomal, 16S; D014157:Transcription Factors; D004247:DNA