URF6, Last Unidentified Reading Frame of Human mtDNA, Codes for an NADH Dehydrogenase Subunit

Abstract

The polypeptide encoded in URF6, the last unassigned reading frame of human mitochondrial DNA, has been identified with antibodies to peptides predicted from the DNA sequence. Antibodies prepared against highly purified respiratory chain NADH dehydrogenase from beef heart or against the cytoplasmically synthesized 49-kilodalton iron-sulfur subunit isolated from this enzyme complex, when added to a deoxycholate or a Triton X-100 mitochondrial lysate of HeLa cells, specifically precipitated the URF6 product together with the six other URF products previously identified as subunits of NADH dehydrogenase. These results strongly point to the URF6 product as being another subunit of this enzyme complex. Thus, almost 60% of the protein coding capacity of mammalian mitochondrial DNA is utilized for the assembly of the first enzyme complex of the respiratory chain. The absence of such information in yeast mitochondrial DNA dramatizes the variability in gene content of different mitochondrial genomes.

Photolabelling of a mitochondrially encoded subunit of NADH dehydrogenase with [3H]dihydrorotenone

“…The subsequent assignment of this and other mitochondrial gene products to NADH dehydrogenase [1,2] has enabled us to identify this protein directly in the purified bovine heart enzyme by immunoblotting. The undecapeptide used as antigen differs by only three amino acids from the equivalent bovine sequence [14,15].…”

“…Seven of the reading frames of the mammalian mitochondrial genome have been shown recently to encode subunits of the respiratory chain NADH dehydrogenase [1,2]. The putative amino acid sequences of these proteins are available for several species, not only mammalian (e.g.…”

Complementary DNA sequences of two 14.5 kDa subunits of NADH:ubiquinone oxidoreductase from bovine heart mitochondria Completion of the primary structure of the complex?

“…Seven of them are encoded in mitochondrial DNA [3][4][5][6], and the remaining subunits are nuclear gene products that are synthesised in the cytoplasm and are imported into the organelle. The sequences of 32 of the nuclear-encoded sttbunits have been described previously [2,[7][8][9][10][11][12][13][14][15][16][17], and they have given many clues about the topograpl~y of the enzyme, and about the functions of some of' the subunits [1].…”

Effects of the Xenopus laevis mitochondrial single‐stranded DNA‐binding protein on the activity of DNA polymerase γ

“…Now that all the open reading frames of the vertebrate mitochondrial genome have been assigned [4], it is clear that the proteins responsible for the replication of the mtDNA are nuclearly encoded. Various enzymes that are probably involved in this process have been identified [5-121 (and references therein).…”

“…A peculiar property of this replication system is that the elongation of the newly synthesized H strand can be stopped at precise sites yielding D loops of various but definite lengths with frequencies usually higher than expected for classical replicative forms (30 -70% in Xenopus hevis oocytes [2]). If the elongation goes through the putative stop signals [3], when two-thirds of the parental H strand are displaced, the origin of light-strand replication becomes exposed and the synthesis of L strand begins in a direction opposite to that of H-strand replication until the formation of two full duplex daughter molecules [l].Now that all the open reading frames of the vertebrate mitochondrial genome have been assigned [4], it is clear that the proteins responsible for the replication of the mtDNA are nuclearly encoded. Various enzymes that are probably involved in this process have been identified [5-121 (and references therein).…”