Sequence and positional requirements for DNA sites in a mu transpososome.
Transposition of bacteriophage Mu uses two DNA cleavage sites and six transposase recognition sites, with each recognition site divided into two half-sites. The recognition sites can activate transposition of non-Mu DNA sequences if a complete set of Mu sequences is not available. We have analyzed 18 sequences from a non-Mu DNA molecule, selected in a functional assay for the ability to be transposed by MuA transposase. These sequences are remarkably diverse. Nonetheless, when viewed as a group they resemble a Mu DNA end, with a cleavage site and a single recognition site. Analysis of these "pseudo-Mu ends" indicates that most positions in the cleavage and recognition sites contribute sequence-specific information that helps drive transposition, though only the strongest contributors are apparent from mutagenesis data. The sequence analysis also suggests variability in the alignment of recognition half-sites. Transposition assays of specifically designed DNA substrates support the conclusion that the transposition machinery is flexible enough to permit variability in half-site spacing and also perhaps variability in the placement of the recognition site with respect to the cleavage site. This variability causes only local perturbations in the protein-DNA complex, as indicated by experiments in which altered and unaltered DNA substrates are paired.
Goldhaber-Gordon
Ilana
I
Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Early
Michael H
MH
Gray
Matthew K
MK
Baker
Tania A
TA
D010584:Bacteriophage phi X 174 / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004251:DNA Transposable Elements; D017353:Gene Deletion; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012689:Sequence Homology, Nucleic Acid; D019895:Transposases / Q000737:chemistry* / Q000378:metabolism
D004251:DNA Transposable Elements; D004247:DNA; C114697:mu transposase; D019895:Transposases