The two single-strand cleavages at each end of Tn10 occur in a specific order during transposition.
During Tn10 transposition, the element is excised from the donor site by double-strand cleavages at the two transposon ends. Double-strand cleavage is a central step in the nonreplicative transposition reaction of many transposons in both prokaryotes and eukaryotes. Evidence is presented to show that the Tn10 double-strand cut is made by an ordered, sequential cleavage of the two strands. The transferred strand is cut first, and then the nontransferred strand is cleaved. The single-strand nicked intermediate is seen to accumulate when Mn2+ is substituted for Mg2+ in the reaction or when certain mutant transposases are used. The fact that the transferred strand is cleaved before the non-transferred strand implies that the order of strand cleavages is not the determining factor that precludes a replicative mechanism of transposition.
Bolland
S
S
Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, USA.
Kleckner
N
N
D004261:DNA Replication; D004251:DNA Transposable Elements; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D019811:Hydroxylamine; D006898:Hydroxylamines; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D016296:Mutagenesis; D009153:Mutagens; D009713:Nucleotidyltransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010957:Plasmids; D019895:Transposases
D004251:DNA Transposable Elements; D006898:Hydroxylamines; D009153:Mutagens; D019811:Hydroxylamine; D008345:Manganese; D009713:Nucleotidyltransferases; D019895:Transposases