A New Mitochondrial Topoisomerase from Rat Liver That Catenates DNA

Abstract

An understanding of the detailed mechanism of mtDNA replication requires studies at the enzyme level. An mtDNA polymerase was first isolated by Meyer and Simpson (1968 Meyer and Simpson (1970) and by Kalf and Ch’ih (1968). Fairfield et al. (1979) have isolated a nicking-closing (N-C) enzyme from rat liver and L-cell mitochondria. This type-I topoisomerase is capable of generating swivels.

Type-II DNA topoisomerases generate transient double-strand breaks and permit a double-stranded segment of DNA to pass through the break prior to resealing (Brown and Cozzarelli 1979; Liu et al. 1980; Mizuuchi et al. 1980). These enzymes not only can relax supercoiled DNA, possibly acting as swivelases, but also can reversibly generate catenanes and knots from closed circles. (Type-I enzymes can also generate catenanes, but at least one of the circles must be nicked [Tse and Wang 1980; Brown and Cozzarelli 1981].) DNA gyrase, a type-II topoisomerase, can also catalyze supercoiling. Such action could periodically introduce negative superturns into replicating mtDNA, keeping the molecule in an even greater energetically favorable state for unwinding W-C turns than could the relaxation induced by the N-C enzyme.

Evidence for an mtDNA gyrase has been obtained (Castora and Simpson 1979; F. J. Castora et al., in prep.). Using intact mitochondria capable of DNA replication (Parsons and Simpson 1967Parsons and Simpson 1973; Karol and Simpson 1968), we showed that in the presence of gyrase inhibitors, i.e., novobiocin. coumermycin, nalidixic acid, and oxolinic acid, (1) incorporation of [³H]dATP into mtDNA is inhibited, (2) incorporation into supercoiled DNA is inhibited...