The terminal protein of a linear mitochondrial plasmid is encoded in the N-terminus of the DNA polymerase gene in white-rot fungus Pleurotus ostreatus

The gene structure and expression of the linear mitochondrial plasmids of t he white-rot fungus Pleurotus ostreatus, pMLP1 and pMLP2, were analyzed. Cl eavage by proteinase K and exonucleases indicated that the 5' ends of pMLP1 and pMLP2 DNAs were associated with terminal proteins. Nucleotide sequenci ng of the entire pMLP1 DNA revealed that it consists of 9,879 bp with termi nal inverted repeat (TIR) sequences of 381 bp. The end sequence of TIR in p MLP1 is 3'-CCCCC-5', similar to those of Escherichia coli phage PRD1. The p MLP1 plasmid harbors two long open reading frames (ORF1 and ORF2) and at le ast one minor ORF (mORF1). The deduced product of ORF1 is homologous to RNA polymerases of yeast mitochondria and several bacteriophages, whereas that of ORF2 is homologous to the protein-primed DNA polymerases of family B ty pe. The mORF1 encodes a highly basic protein, most likely a TIR-binding pro tein, with no apparent sequence homology in the database. Expression of the predicted gene products from pMLP1 in mitochondria was demonstrated by Wes tern blot analysis using antibodies against various expressed regions of pM LP1 ORFs. A plasmid-free strain, generated by curing with ethidium bromide, did not express any of these gene products. Terminal proteins of 70 kDa (T P1) and 73 kDa (TP2) were identified from pMLP1 and pMLP2, respectively. We stern blot analysis indicated that TP1 was generated from the N-terminal ha lf of the full-length product of ORF2 encoding a putative DNA polymerase.