Authors
Alagha, T., Arin, A., Vangos, N., Goodey-Parfitt, H., Ngo, H. N., Dau, N. N., Nguyen, M. H., Legal, T., Cianfrocco, M. A., Bui, K. H.
Abstract
Microtubules are cytoskeletal filaments that are involved in intracellular transport, cell division, and motility. Despite their biological importance, determining their high-resolution structures via cryo-electron microscopy remains a significant technical challenge due to their polymorphisms and pseudo-helical assembly. Current processing workflows are complex, often requiring the integration of multiple software packages and custom scripts, which creates a steep learning curve for many research groups. To address these limitations, we introduce CsMT, a streamlined workflow implemented entirely within the CryoSPARC environment and using synthetic references. CsMT simplifies microtubule reconstruction by utilizing a novel protofilament-pair classification approach, which effectively handles the inherent pseudo-symmetry and structural heterogeneity of microtubules with minimal manual intervention. Our workflow is versatile, capable of processing both undecorated and decorated microtubules while accurately determining seams and performing high-resolution refinement. We demonstrate the efficacy of this workflow by achieving a 2.3 and 2.7 [A] resolution reconstruction of homotypic and heterotypic maps of undecorated microtubules, matching the best-resolved microtubule structures in the field. By unifying the pipeline into a single, portable workflow, CsMT enhances reproducibility and accessibility, empowering more laboratories to explore the structural biology of microtubules and associated proteins, yielding new insights into their function.
Preprint server:
bioRxiv
The authors list and abstract were imported from bioRxiv on 05 Aug 2026.
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