Yeast mtDNA is compacted into nucleoprotein structures called mitochondrial nucleoids (mt-nucleoids). The principal mediators of nucleoid formation are mitochondrial high-mobility group (HMG)-box containing (mtHMG) proteins. Although these proteins are some of the fastest evolving components of mt-nucleoids, it is not known whether the divergence of mtHMG proteins on the level of their amino acid sequences is accompanied by diversification of their biochemical properties. In the present study we performed a comparative biochemical analysis of yeast mtHMG proteins from Saccharomyces cerevisiae (ScAbf2p), Yarrowia lipolytica (YlMhb1p) and Candida parapsilosis (CpGcf1p). We found that all three proteins exhibit relatively weak binding to intact dsDNA. In fact, ScAbf2p and YlMhb1p bind quantitatively to this substrate only at very high protein to DNA ratios and CpGcf1p shows only negligible binding to dsDNA. In contrast, the proteins exhibit much higher preference for recombination intermediates such as Holliday junctions (HJ) and replication forks (RF). Therefore, we hypothesize that the roles of the yeast mtHMG proteins in maintenance and compaction of mtDNA in vivo are in large part mediated by their binding to recombination/replication intermediates. We also speculate that the distinct biochemical properties of CpGcf1p may represent one of the prerequisites for frequent evolutionary tinkering with the form of the mitochondrial genome in the CTG-clade of hemiascomycetous yeast species.

Department of Biology Masaryk University Kamenice 5 A7 Brno 625 00 Czech Republic International Clinical Research Center St Anne's University Hospital in Brno Brno 60200 Czech Republic

Department of Biology Masaryk University Kamenice 5 A7 Brno 625 00 Czech Republic International Clinical Research Center St Anne's University Hospital in Brno Brno 60200 Czech Republic National Center for Biomolecular Research Masaryk University Kamenice 5 A7 Brno 625 00 Czech Republic

Departments of Genetics and Biochemistry Comenius University in Bratislava Faculty of Natural Sciences Mlynska dolina Ilkovicova 6 842 15 Bratislava Slovak Republic

Lineberger Comprehensive Cancer Center University of North Carolina at Chapel Hill NC 27599 U S A

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