Vemu, A;
Atherton, J;
Spector, JO;
Szyk, A;
Moores, CA;
Roll-Mecak, A;
(2016)
Structure and Dynamics of Single-isoform Recombinant Neuronal Human Tubulin.
Journal of Biological Chemistry
, 291
(25)
pp. 12907-12915.
10.1074/jbc.C116.731133.
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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. Although 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 homogeneous 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 post-translationally unmodified human α1A/βIII microtubules shows overall similarity to that of heterogeneous brain microtubules, but it is distinguished by subtle differences at polymerization interfaces, which are hot spots for sequence divergence between tubulin isoforms. In vitro dynamics assays show that, like mosaic brain microtubules, recombinant homogeneous microtubules undergo dynamic instability, but they polymerize slower and have fewer catastrophes. 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 homogeneous 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 toward uncovering how tubulin genetic and chemical diversity is exploited to modulate intrinsic microtubule dynamics.
| Type: | Article |
|---|---|
| Title: | Structure and Dynamics of Single-isoform Recombinant Neuronal Human Tubulin |
| Location: | United States |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1074/jbc.C116.731133 |
| Publisher version: | http://doi.org/10.1074/jbc.C116.731133 |
| Language: | English |
| Additional information: | © 2016 by The American Society for Biochemistry and Molecular Biology, Inc. Author's Choice—Final version free via Creative Commons CC-BY license (http://creativecommons.org/licenses/by/4.0). |
| Keywords: | cryo-electron microscopy, cytoskeleton, microscopy, microtubule, tubulin, dynamic instability, engineered tubulin, microtubule dynamics, recombinant tubulin, tubulin isoform, microtubule structure, cryo-EM, post-translational modification |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Structural and Molecular Biology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10073663 |
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