Tn5 transposase loops DNA in the absence of Tn5 transposon end sequences.
Transposases mediate transposition first by binding specific DNA end sequences that define a transposable element and then by organizing protein and DNA into a highly structured and stable nucleoprotein 'synaptic' complex. Synaptic complex assembly is a central checkpoint in many transposition mechanisms. The Tn5 synaptic complex contains two Tn5 transposase subunits and two Tn5 transposon end sequences, exhibits extensive protein-end sequence DNA contacts and is the node of a DNA loop. Using single-molecule and bulk biochemical approaches, we found that Tn5 transposase assembles a stable nucleoprotein complex in the absence of Tn5 transposon end sequences. Surprisingly, this end sequence-independent complex has structural similarities to the synaptic complex. This complex is the node of a DNA loop; transposase dimerization and DNA specificity mutants affect its assembly; and it likely has the same number of proteins and DNA molecules as the synaptic complex. Furthermore, our results indicate that Tn5 transposase preferentially binds and loops a subset of non-Tn5 end sequences. Assembly of end sequence-independent nucleoprotein complexes likely plays a role in the in vivo downregulation of transposition and the cis-transposition bias of many bacterial transposases.
Adams
Christian D
CD
Department of Biochemistry, University of Wisconsin at Madison, 433 Babcock Drive, WI 53706, USA.
Schnurr
Bernhard
B
Skoko
Dunja
D
Marko
John F
JF
Reznikoff
William S
WS
D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004251:DNA Transposable Elements / Q000235:genetics*; D019281:Dimerization; D024202:Electrophoretic Mobility Shift Assay; D008954:Models, Biological; D008958:Models, Molecular; D009154:Mutation; D009690:Nucleic Acid Conformation; D009698:Nucleoproteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D019895:Transposases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D004251:DNA Transposable Elements; D009698:Nucleoproteins; C473503:Tn5 transposase; D004247:DNA; D019895:Transposases