title: Phosphatidylcholine and phosphatidylethanolamine enhance the activity of the mammalian mitochondrial endonuclease in vitro.
abstract: The purified endonuclease of bovine heart mitochondria extensively degrades a variety of DNA templates in vitro but shows a remarkably strong preference to nick within one specific evolutionarily conserved sequence block of 12 consecutive guanine residues which resides just upstream from the heavy strand origin of mitochondrial DNA replication (Low, R. L., Cummings, O. W., and King, T. C. (1987) J. Biol. Chem. 262, 16164-16170). If the enzyme serves to provide an important nicking function at this site in vivo, then mitochondrial factors likely exist which further enhance the enzyme's recognition of this locus and prevent cleavage at other less favored sites. In this study, we report that specific membrane phospholipids appear to exert such effects in vitro. In standard endonuclease assays, low levels of phosphatidylcholine or phosphatidylethanolamine (0.5 mM) stimulate the purified enzyme activity 10-20-fold. However, at moderate levels (20-40 mM), these phospholipids largely inhibit widespread degradation of duplex DNA while still allowing site-specific nicking at the conserved guanine target in the mitochondrial genome. These findings suggest that an interaction of the endonuclease with major lipid components of the inner membrane could be an important determinant of the enzyme's specificity for mitochondrial DNA.
authors.affiliation: Department of Pathology, University of Colorado Health Sciences Center, Denver 80262.
authors.forename: W A
authors.initials: WA
authors.lastname: Parks
authors.forename: C L
authors.initials: CL
authors.lastname: Couper
authors.forename: R L
authors.initials: RL
authors.lastname: Low
mesh_terms: D000818:Animals; D002417:Cattle; D004272:DNA, Mitochondrial / Q000378:metabolism; D004720:Endonucleases / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008929:Mitochondria, Heart / Q000201:enzymology*; D010713:Phosphatidylcholines / Q000494:pharmacology*; D010714:Phosphatidylethanolamines / Q000494:pharmacology*; D013379:Substrate Specificity
chemical_list: D004272:DNA, Mitochondrial; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D004720:Endonucleases