Cryo-EM imaging of the catalytic subunit of the DNA-dependent protein kinase.

The DNA-dependent protein kinase (DNA-PK) plays an important role in mammalian DNA double-strand break repair and immunoglobulin gene rearrangement. The DNA-PK holoenzyme is activated by assembly at DNA ends and is comprised of DNA-PKcs, a 460 kDa protein kinase catalytic subunit, and Ku, a 70 kDa/80 kDa heterodimeric DNA-targeting component. We have solved the three-dimensional structure of DNA-PKcs to approximately 21 A resolution by analytically combining images of nearly 9500 individual particles extracted from cryo-electron micrographs. The DNA-PKcs protein has an open, pseudo 2-fold symmetric structure with a gap separating a crown-shaped top from a rounded base. Columns of density are observed to protrude into the gap from both the crown and the base. Measurements of the enclosed volume indicate that the interior of the protein is largely hollow. The structure of DNA-PKcs suggests that its association with DNA may involve the internalization of double-stranded ends.

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