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RESEARCH PAPER

3D reconstruction of the Mu transposase and the Type 1 transpososome: a structural framework for Mu DNA transposition

¹ Department of Biochemistry, University of Western Ontario, London, Ontario N6A 5C1, Canada; ² Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G 2M9, Canada; ³ Department of Biochemistry and Molecular Biology, and Department of Microbiology and Infectious Diseases, University of Calgary, Calgary, Alberta T2N 4N1, Canada

Mu DNA transposition proceeds through a series of higher-order nucleoprotein complexes called transpososomes. The structural core of the transpososome is a tetramer of the transposase, Mu A, bound to the two transposon ends. High-resolution structural analysis of the intact transposase and the transpososome has not been successful to date. Here we report the structure of Mu A at 16-Å and the Type 1 transpososome at 34-Å resolution, by 3D reconstruction of images obtained by scanning transmission electron microscopy (STEM) at cryo-temperatures. Electron spectroscopic imaging (ESI) of the DNA-phosphorus was performed in conjunction with the structural investigation to derive the path of the DNA through the transpososome and to define the DNA-binding surface in the transposase. Our model of the transpososome fits well with the accumulated biochemical literature for this intricate transposition system, and lays a structural foundation for biochemical function, including catalysis in trans and the complex circuit of macromolecular interactions underlying Mu DNA transposition.

[Keywords: DNA transposition; transpososome; transposase; site-specific recombination; 3D reconstruction; electron spectroscopic imaging; scanning transmission electron microscopy]

Received December 20, 2004; revised version accepted February 2, 2005.

Article published online ahead of print. Article and publication date are at http://www.genesdev.org/cgi/doi/10.1101/gad.1291405.

⁴ Present address: National Microbiology Laboratory, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada.

⁵ Corresponding author.

E-MAIL chaconas{at}ucalgary.ca; FAX (403) 270-2772.

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