Rad5l protein of Saccharomyces cerevisiae is a structural homolog of the Escherichia coli recombination enzyme RecA. In yeast, the Rad5l protein is required for mitotic and meiotic recombination and for repair of doublestrand breaks in DNA. We have used antibodies raised against the homologous human protein, HsRad51, expressed

A general model is proposed for genetic recombination. Its essential new feature is the hypothesis that recombination is initiated by a single-strand (or asymmetric) transfer, which may, after isomerization, become a two-strand (or symmetric) exchange. The likelihood of this transition from asymmetric to symmetric strand exchange determines certain characteristic features of recombination in any particular organism.In eukaryotes the segregation of an allele pair among the four products of meiosis occasionally departs from the usual 2:2 ratio while the segregation of well-separated markers on either side remains normal (i.e., 2:2). Frequently the flanking markers are recombined. This pattern, derived largely from genetic studies in fungi, probably means that the region of aberrant segregation is the actual site of chemical interaction between two recombining DNA molecules. Genetic analysis in fungi and prokaryotes, considerations of DNA structure, and the recognition of a number of enzymic mechanisms of DNA chemistry have made it possible to formulate rather specific hypotheses about molecular aspects of recombination, as we do in this paper (see refs. 1 and 2 for reviews). The Holliday modelIn 1964 Holliday proposed a model to explain recombination and aberrant segregation in fungi (3). Two homologous DNA duplexes, each corresponding to one of the four chromatids present at meiosis, undergo single-strand breakage and exchange single strands, forming heteroduplex DNA symmetrically for a limited distance as depicted in Fig. 1 (p,p in Fig. 1) leaves the flanking arms in the parental configuration, whereas cleavage of the other pair of strands (r,r) yields products with the flanking arms in the recombinant configuration.Recently, Sigal and Alberts (4) constructed a molecular model of the Holliday structure and found that it can be built with satisfactory bond lengths and angles, with no bases unpaired. The cross connection can move up or down by rotation of both duplexes in the same sense, a process that could be driven by rotary diffusion (5). Migration of the cross connection extends the region of heteroduplex DNA symmetrically on both chromatids, giving the postulated recombination intermediate (Fig. 1). Heteroduplex DNA in one versus two chromatidsThe equal formation of heteroduplex DNA on both chromatids, which is an intrinsic feature of the Holliday structure, should be reflected in the genetic data. The relevant evidence is mixed. The occurrence of aberrant 4:4 segregations in Sordaria indicates that heteroduplex DNA can form at the same site on both chromatids (6). At the b2 locus in Ascobolus, Leblon and Rossignol (7) observed that the ratios of 6:2, 5:3, and aberrant 4:4 segregations were in good agreement with the hypothesis that heteroduplex DNA usually forms on both chromatids. In contrast, other studies are most simply interpreted to mean that heteroduplex DNA usually forms on only one chromatid. At the arg4 locus in yeast the frequency of intragenic two-strand (i.e., two-chromatid) double e...

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Homologous pairing and strand exchange, which are catalyzed by Escherichia coli RecA protein, are central to homologous recombination. Homologs of this protein are found in eukaryotes; however, little has been reported on the recombinase activities of the mammalian homologs, including the human protein, denoted HsRad51. For the studies described here, we purified HsRad51 from E. coli. Although the activities of HsRad51 and RecA were qualitatively similar in the presence of ATP, there were also striking differences. The stoichiometry of binding to DNA and the rate of renaturation of complementary strands were similar for the two proteins, but rates of ATP hydrolysis, homologous pairing, and subsequent strand exchange promoted by HsRad51 were less than 1 ⁄10 those of RecA. In addition, HsRad51 bound ␥-thio-ATP and formed stable presynaptic complexes that promoted renaturation as rapidly as RecA, but the recombinant human protein catalyzed neither strand exchange nor homologous pairing of a single strand with duplex DNA in the presence of the ATP analog. By contrast, RecA promoted both of the latter reactions in control experiments. These observations suggest that among RecA-like proteins, HsRad51 may be a variant in which homologous pairing and strand exchange are more closely linked to the hydrolysis of ATP.

The ␤ protein of bacteriophage acts in homologous genetic recombination by catalyzing the annealing of complementary single-stranded DNA produced by the exonuclease. It has been shown that the ␤ protein binds to the products of the annealing reaction more tightly than to the initial substrates. We find that ␤ protein exists in three structural states. In the absence of DNA, ␤ protein forms inactive rings with Ϸ12 subunits. The active form of the ␤ protein in the presence of oligonucleotides or single-stranded DNA is a ring, composed of Ϸ15-18 subunits. The doublestranded products of the annealing reaction catalyzed by the rings are bound by ␤ protein in a left-handed helical structure, which protects the products from nucleolytic degradation. These observations suggest structural homology for a family of proteins, including the phage P22 erf, the bacterial RecT, and the eukaryotic Rad52 proteins, all of which are involved in homologous recombination.