CHARACTERIZATION OF A DNA PAIRING ACTIVITY COPURIFYING WITH DNA-LIGASE IN A PARTIALLY PURIFIED EXTRACT FROM RAT TESTIS

Rat testicular nuclear extracts were fractionated sequentially on phos phocellulose, heparin-agarose and ssDNA-cellulose columns, in order to isolate and characterise a strand-transfer activity from a mammalian meiotic tissue. A partially purified fraction, eluting at 0.6 M KCI fr om ssDNA-cellulose column, catalyzed the formation of two classes of p roducts migrating slowly on an agarose gel. The formation of one of th ese classes of products - the aggregates - was dependent on the presen ce of both the substrates (M13mp19 RF III and M13mp19 ssDNA) and on ho mology. The presence of ATP was essential for the formation of aggrega tes, though its hydrolysis was not required. EM analysis of the produc ts indicated the presence of structures which resembled paired DNA mol ecules: duplex-duplex paired (Y-shaped and ds-ds paired structures) an d ss-ds paired (duplex DNA paired with the single-stranded DNA) struct ures, indicating the presence of a pairing protein in the fraction. Ho wever, alpha- and sigma-structures were not observed, The other class of products, seen as discrete bands, were identified biochemically and by electron microscopy as ligated products. A DNA ligase-adenylate ad duct of molecular weight 100 kDa was formed by the fraction. Both 5' t o 3' and 3' to 5' exonucleases were absent and hence did not contribut e to the formation of the products.