THE 3 CHEMICAL STEPS OF TN10 IS10 TRANSPOSITION INVOLVE REPEATED UTILIZATION OF A SINGLE ACTIVE-SITE/

Nonreplicative transposition by Tn10/IS10 involves three chemical step s at each transposon end: cleavage of the two strands plus joining of one strand to target DNA. These steps occur within a synaptic complex comprising two transposon ends and monomers of IS10 transposase. We re port four transposase mutations that individually abolish each of the three chemical steps without affecting the synaptic complex. We conclu de that a single constellation of residues, the ''active site,'' direc tly catalyzes each of the three steps. Analyses of reactions containin g mixtures of wild-type and catalysis-defective transposases indicate that a single transposase monomer at each end catalyzes the cleavage o f two strands and that strand transfer is carried out by the same mono mers that previously catalyzed cleavage. These and other data suggest that one active site unit carries out all three reactions in successio n at one transposon end.