The human mitochondrial transcription termination factor (mTERF) is a multizipper protein but binds to DNA as a monomer, with evidence pointing to intramolecular leucine zipper interactions

Abstract

The human mitochondrial transcription termination factor (mTERF) cDNA has been cloned and expressed in vitro, and two alternative precursors of the protein have been imported into isolated mitochondria and processed to the mature protein. The precursors contain a mitochondrial targeting sequence, and the mature mTERF (342 residues) exhibits three leucine zippers, of which one is bipartite, and two widely spaced basic domains. The in vitro synthesized mature protein has the expected specific binding capacity for a double‐stranded oligonucleotide containing the tridecamer sequence required for directing termination, and produces a DNase I footprint very similar to that produced by the natural protein. However, in contrast to the latter, it lacks transcription termination‐promoting activity in an in vitro system, pointing to another component(s) being required for making mTERF termination‐competent. A detailed structure–function analysis of the recombinant protein and mutagenized versions of it by band shift assays has demonstrated that both basic domains and the three leucine zipper motifs are necessary for DNA binding. Furthermore, a variety of tests have shown that both the recombinant and the natural mTERF bind to DNA as a monomer, arguing against a dimerization role for the leucine zippers, and rather pointing, together with the results of mutagenesis experiments, to intramolecular leucine zipper interactions being required to bring the two basic domains in close register with the mTERF target DNA sequence.

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

“…One is a monomeric form, exhibiting DNA binding and transcriptiontermination activities, as expected from previous results (9), whereas the other is a novel high molecular weight complex lacking DNA binding activity. The evidence obtained has strongly suggested that the novel complex form of mTERF is a homotrimer.…”

“…Both types of motifs are essential for binding of mTERF to its DNA target sequence, and, consequently, for promoting transcription termination (9). Despite leucine zippers being typical protein-protein interaction motifs, it has been clearly demonstrated that mTERF binds DNA as a monomer (9).…”

“…Despite leucine zippers being typical protein-protein interaction motifs, it has been clearly demonstrated that mTERF binds DNA as a monomer (9). This has led to the suggestion that mTERF acquires DNA binding activity by means of intramolecular leucine-zipper interactions, required to bring the two basic DNA-binding domains together (9).…”

pTAC2, -6, and -12 Are Components of the Transcriptionally Active Plastid Chromosome That Are Required for Plastid Gene Expression

“…Eight additional proteins contain DNA/RNA binding domains, such as the PPR, SMR, SAP, OB fold, S1, KOW, NGN, mTERF, or single-stranded DNA (ssDNA) binding motifs (Table 2). These motifs are found in proteins involved in chromatin organization, DNA repair, transcription, RNA processing, or translation (Boni et al, 1991;Kyrpides et al, 1996;Fernandez-Silva et al, 1997;Draper and Reynaldo, 1999;Moreira and Philippe, 1999;Aravind and Koonin, 2000;Lurin et al, 2004). Furthermore, pTAC1 and pTAC11 exhibit striking similarities to p24 proteins, which are members of the Whirly proteins of nuclear transcription factors (Desveaux et al, 2002(Desveaux et al, , 2004.…”

Cluster analysis of gene expression dynamics

“…The cluster includes receptor tyrosine kinase-like orphan receptor 2, TRAF-binding protein domain, and Deathassociated protein kinase. The cluster also includes the transcription termination factor-like protein, which plays a central role in the control of rRNA and mRNA synthesis in mammalian mitochondria (28), and DKFZP586G1122 protein, which has unknown function but has strong homology with murine zinc finger protein Hzf expressed in hematopoiesis.…”