title: 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.
authors.affiliation: Division of Biology, California Institute of Technology, Pasadena 91125, USA.
authors.forename: P
authors.initials: P
authors.lastname: Fernandez-Silva
authors.forename: F
authors.initials: F
authors.lastname: Martinez-Azorin
authors.forename: V
authors.initials: V
authors.lastname: Micol
authors.forename: G
authors.initials: G
authors.lastname: Attardi
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D018983:DNA Footprinting; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007460:Iodoacetamide / Q000378:metabolism; D016350:Leucine Zippers; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism*; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D017354:Point Mutation; D011489:Protein Denaturation; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000378:metabolism; D017421:Sequence Analysis; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014357:Trypsin / Q000378:metabolism
chemical_list: D050976:Basic-Leucine Zipper Transcription Factors; C105178:MTERF1 protein, human; D024101:Mitochondrial Proteins; D011498:Protein Precursors; D011994:Recombinant Proteins; D014157:Transcription Factors; D004247:DNA; D014357:Trypsin; D007460:Iodoacetamide