A microtubule RELION-based pipeline for cryo-EM image processing

A. D Cook, S. W Manka, S. Wang, C. A Moores, Joseph Michael Atherton

Research output: Contribution to journalArticlepeer-review

28 Citations (Scopus)


Microtubules are polar filaments built from αβ-tubulin heterodimers that exhibit a range of architectures in vitro and in vivo. Tubulin heterodimers are arranged helically in the microtubule wall but many physiologically relevant architectures exhibit a break in helical symmetry known as the seam. Noisy 2D cryo-electron microscopy projection images of pseudo-helical microtubules therefore depict distinct but highly similar views owing to the high structural similarity of α- and β-tubulin. The determination of the αβ-tubulin register and seam location during image processing is essential for alignment accuracy that enables determination of biologically relevant structures. Here we present a pipeline designed for image processing and high-resolution reconstruction of cryo-electron microscopy microtubule datasets, based in the popular and user-friendly RELION image-processing package, Microtubule RELION-based Pipeline (MiRP). The pipeline uses a combination of supervised classification and prior knowledge about geometric lattice constraints in microtubules to accurately determine microtubule architecture and seam location. The presented method is fast and semi-automated, producing near-atomic resolution reconstructions with test datasets that contain a range of microtubule architectures and binding proteins.

Original languageEnglish
Article number107402
JournalJournal of Structural Biology
Issue number1
Publication statusPublished - 2020


  • 3D reconstruction
  • Cryo-EM
  • Microtubule
  • Pseudo-helical symmetry


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