BIROn - Birkbeck Institutional Research Online

    Structure and dynamics of single-isoform recombinant Neuronal Human Tubulin

    Vemu, A. and Atherton, Joe and Spector, J.O. and Szyk, A. and Moores, Carolyn A. and Roll-Mecak, A. (2016) Structure and dynamics of single-isoform recombinant Neuronal Human Tubulin. Journal of Biological Chemistry 291 (25), pp. 12907-12915. ISSN 0021-9258.

    [img]
    Preview
    Text
    15034.pdf - Author's Accepted Manuscript

    Download (9MB) | Preview
    [img]
    Preview
    Text (Table1)
    15034a.pdf - Supplemental Material

    Download (83kB) | Preview
    [img] Video (Movie1)
    15034b.avi - Supplemental Material

    Download (57MB)
    [img] Video (Movie2)
    15034c.avi - Supplemental Material

    Download (46MB)
    [img] Video (Movie3)
    15034d.avi - Supplemental Material

    Download (2MB)

    Abstract

    Microtubules are polymers that cycle stochastically between polymerization and depolymerization i.e., they exhibit 'dynamic instability'. This behavior is crucial for cell division, motility and differentiation. While studies in the last decade have made fundamental breakthroughs in our understanding of how cellular effectors modulate microtubule dynamics, analysis of the relationship between tubulin sequence, structure and dynamics has been held back by a lack of dynamics measurements with and structural characterization of homogenous, isotypically pure, engineered tubulin. Here we report for the first time the cryo-EM structure and in vitro dynamics parameters of recombinant isotypically pure human tubulin. α1A/βIII is a purely neuronal tubulin isoform. The 4.2 Å structure of unmodified human α1A/βIII microtubules shows overall similarity to that of heterogeneous brain microtubules, but is distinguished by subtle differences at polymerization interfaces, which are hotspots for sequence divergence between tubulin isoforms. In vitro dynamics assays show that, like mosaic brain microtubules, recombinant homogenous microtubules undergo dynamic instability but they polymerize slower and catastrophe less frequently. Interestingly, we find that epitaxial growth of α1A/βIII microtubules from heterogeneous brain seeds is inefficient, but can be fully rescued by incorporating as little as 5% of brain tubulin into the homogenous α1A/βIII lattice. Our study establishes a system to examine the structure and dynamics of mammalian microtubules with well-defined tubulin species and is a first and necessary step towards uncovering how tubulin genetic and chemical diversity is exploited to modulate intrinsic microtubule dynamics.

    Metadata

    Item Type: Article
    Keyword(s) / Subject(s): cryo-electron microscopy, cytoskeleton, microscopy, microtubule, dynamics, recombinant tubulin, tubulin isoform tubulin, dynamic instability, engineered tubulin, microtubule
    School: Birkbeck Schools and Departments > School of Science > Biological Sciences
    Research Centre: Structural Molecular Biology, Institute of (ISMB)
    Depositing User: Administrator
    Date Deposited: 27 Apr 2016 09:17
    Last Modified: 13 Dec 2019 19:46
    URI: http://eprints.bbk.ac.uk/id/eprint/15034

    Statistics

    Downloads
    Activity Overview
    247Downloads
    129Hits

    Additional statistics are available via IRStats2.

    Archive Staff Only (login required)

    Edit/View Item Edit/View Item