Intracellular Localization of Rat‐Liver Uracil‐DNA Glycosylase

Abstract

Most of the uracil-DNA glycosylase of the rat liver cell is located in chromatin; there is, however, some activity in the nuclear sap and in the cytoplasm. The chromatin uracil-DNA glycosylase has been purified; the preparation is devoid of endonuclease and exonuclease activities ; the enzyme does not need divalent cations, has a broad optimum pH around 8, is strongly inhibitcd by increasing ionic strength and free uracil.The apparent K, is independent of the strandedness of the DNA substrate containing uracil, but V is slightly higher with the single-stranded substrate. The frequency of uracil substitution in the double-stranded DNA influences the kinetic parameters: a higher frequency increases both K, and V.The inhibitory effects of NaCl and free uracil are greater when the substrate is double-stranded rather than single-stranded. It is speculated that, acting either on the D N A or on the enzyme, both oppose the opening of the double helix necessary for the formation of the enzyme-substrate complex.The increased reaction rate with a higher frequency of uracil residues in double-stranded D N A is interpreted as a tendency for the repair enzyme to work in a processive way. It is supposed that pmcessivity also occurs with single-stranded D N A and that it is opposed by both NaCl and free uracil, explaining a greater inhibition when the single-stranded substrate has a higher uracil content.

Ap endonucleases and dna glycosylases that recognize oxidative dna damage

“…Both nuclear and mitochondria1 forms of uracil DNA glycosylases have been found in human KB [Anderson and Friedberg, 19801, WI-38 Sirover, 1981b1, and HeLa [Witwer and Krokan, 19851 cells. Both nuclear and mitochondria1 forms of the enzyme have also been observed in rat liver [Colson and Verly, 1983;Domena and Mosbaugh, 19851. Uracil DNA Glycosylases and DNA Repair E. cofi mutants defective in uracil DNA glycosylase, called ung, have been isolated [Duncan et al, 19781. These mutants exhibit an increased spontaneous mutation rate of the G:C to A:T transition type [Duncan and Miller, 1980;Duncan and Weiss, 1982;Fix and Glickman, 1986, 19871.…”

Nuclear and mitochondrial forms of human uracil-DNA glycosylase are encoded by the same gene

“…The nuclear form of uracil-DNA glycosylase is far more abundant than the mitochondrial form (7,15,16). Transcription of the UNG gene is up-regulated in the late GI-phase, accounting for more than 90% of the uracil-DNA glycosylase activity at the early S-phase (17).…”

Properties of a Recombinant Human Uracil-DNA Glycosylase from the UNG Gene and Evidence that UNG Encodes the Major Uracil-DNA Glycosylase

“…In eukaryotic cells the major fraction of uracil-DNA glycosylase activity is located in the nuclei with a minor fraction in the mitochondria (Anderson & Friedberg, 1980;Gupta & Sirover, 1981;Colson & Verly, 1983; Wittwer & Krokan, 1985). cDNA for human conserved uracil-DNA glycosylase (UNG15) (Olsen et al, 1989) encodes a major human uracil-DNA glycosylase (Slupphaug et al, 1991).…”