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. 2008 Mar 20;27(7):1097–1109. doi: 10.1038/emboj.2008.41

Figure 7.

Figure 7

Mutation of conserved DDD amino acids blocks the catalytic activity of piggyBac transposase in vitro. (A) Mutation of the conserved D's blocks 3′OH nicking. Wild-type and mutant piggyBac transposases were incubated with a piggyBac L-TIR and a target plasmid for 20 min and then displayed on a denaturing acrylamide gel. (B) Mutation of the conserved D's blocks hairpin formation. Wild-type and mutant piggyBac transposases were incubated with a pre-formed piggyBac L-TIR hairpin oligonucleotide and then displayed on a denaturing gel. (C) Mutation of the conserved D's blocks target joining. Wild-type and mutant transposases were incubated with a pre-cleaved piggyBac L-TIR lacking the usual 4 nt overhangs at 5′ transposon end and a target plasmid and then displayed on a native agarose gel. ∞ indicates nucleoprotein complexes formed by transposase binding to the labelled piggyBac L-TIRs.