S. Bolland et N. Kleckner, THE 3 CHEMICAL STEPS OF TN10 IS10 TRANSPOSITION INVOLVE REPEATED UTILIZATION OF A SINGLE ACTIVE-SITE/, Cell, 84(2), 1996, pp. 223-233
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.