SITE-SPECIFIC RECOMBINATION AT RES SITES CONTAINING DNA-BINDING SEQUENCES FOR BOTH TN21 AND TN3 RESOLVASES

Tn3 and gamma delta resolvases catalyse site-specific recombination at res sites from Tn3 but not at Tn21 res sites, Tn21 resolvase has no a ctivity at Tn3 sites and acts only at Tn21 sites. In both Tn3 and Tn21 , res has three binding sites for the cognate resolvases; the cross-ov er site, I; and the accessory sites II and III, from which the bound p roteins may stabilize the synaptic complex by protein-protein interact ions. In this study hybrid I es sites were made by replacing either II or III in the Tn21 res site with the equivalent sequence from Tn3. Pl asmids containing either a hybrid and a wild-type Tn21 yes site, or tw o hybrid sites, were tested for recombination. Relative to the reactio n with two wild-type sites, recombination by Tn21 resolvase was reduce d by replacing II at one res site and it was reduced further by replac ing II at both loci but, in both cases, Tn21 recombination was enhance d by Tn3 or gamma delta resolvases. Very few of the amino acid on the external surface of gamma delta resolvases are conserved in Tn21. More over, mutants of gamma delta resolvase with defective protein-protein interactions also enhanced Tn21 recombination at this hybrid site. The resolvase at II thus seems not to be involved in protein-protein inte ractions and its main role may be to bend the DNA to the required stru cture. The replacement of III in the Tn21 site with Tn3 sequence also reduced recombination by Tn21 resolvase, especially when both loci car ried the alteration but, in contrast to before, Tn3 or gamma delta res olvases now inhibited the Tn21 reaction. Recombination thus seems to r equire identical proteins at I and III, perhaps to allow for protein-p rotein interactions.