Characterization of the TnsD-attTn7 complex that promotes site-specific insertion of Tn7.
The bacterial transposon Tn7 is distinguished by its ability to recognize a specific site called attTn7, and insert just downstream of the highly conserved chromosomal glmS gene. TnsD is one of four transposon-encoded polypeptides (TnsABC+D) required for site-specific insertion of Tn7 into attTn7, and is the target site-selector that binds to a highly conserved sequence in the end of the glmS protein coding region. In this study, we identified important nucleotides within this region that are crucial for TnsD-attTn7 interaction. We also probed the regions of TnsD that interact with attTn7 and found that there are important DNA-binding determinants throughout the entire length of the protein, including an amino-terminal CCCH zinc-finger motif. A key role of TnsD is to recruit the non-sequence specific DNA-binding protein TnsC to attTn7; TnsC also interacts with and controls both the TnsA and TnsB subunits of the Tn7 transposase. TnsC stimulates the binding of TnsD to attTn7 in vivo, and TnsCD and TnsD can also interact in the absence of DNA and localize their interaction domains to the N-terminal region of each protein.
Mitra
Rupak
R
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore MD 21205, USA.
McKenzie
Gregory J
GJ
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore MD 21205, USA.|Current Address: Verenium Corporation. 4955 Directors Place, San Diego, CA 92121, USA.
Yi
Liang
L
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore MD 21205, USA.|Current Address: Laboratory of Host Defense, NIAID/NIH, Bethesda, MD 20892, USA.
Lee
Cherline A
CA
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore MD 21205, USA.|Current Address: Mayo Clinic, 417 Guggenheim Bldg, 200 First St. SW, Rochester, MN 55905, USA.
Craig
Nancy L
NL
Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore MD 21205, USA.