Challenges and opportunities in the high-resolution cryo-EM visualization of microtubules and their binding partners

Nogales, Eva; Kellogg, Elizabeth H.

doi:10.1016/j.sbi.2017.06.003

Cited by 18 publications

(11 citation statements)

References 34 publications

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“…In vivo , it most often comprises 13 parallel protofilaments in which tubulin molecules are arranged head to tail, defining a (fast growing) plus end terminated by β-tubulin and a (slower growing) minus end exposing α-tubulin. As such, it cannot be crystallized but is well suited for cryo-EM studies (reviewed in Downing and Nogales, 1998 ; Manka and Moores, 2018a ; Nogales and Kellogg, 2017 ). Actually, it has fully benefited from the advances of this method, the resolution having improved from 20 to about 3 Å in less than 20 years ( Nogales et al., 1999 ; Zhang et al., 2018 ).…”

Section: Complementary Methods For the Structural Study Of Tubulin Anmentioning

confidence: 99%

The Mechanism of Tubulin Assembly into Microtubules: Insights from Structural Studies

et al. 2020

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Summary Microtubules are cytoskeletal components involved in pivotal eukaryotic functions such as cell division, ciliogenesis, and intracellular trafficking. They assemble from αβ-tubulin heterodimers and disassemble in a process called dynamic instability, which is driven by GTP hydrolysis. Structures of the microtubule and of soluble tubulin have been determined by cryo-EM and by X-ray crystallography, respectively. Altogether, these data define the mechanism of tubulin assembly-disassembly at atomic or near-atomic level. We review here the structural changes that occur during assembly, tubulin switching from a curved conformation in solution to a straight one in the microtubule core. We also present more subtle changes associated with GTP binding, leading to tubulin activation for assembly. Finally, we show how cryo-EM and X-ray crystallography are complementary methods to characterize the interaction of tubulin with proteins involved either in intracellular transport or in microtubule dynamics regulation.

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Section: Complementary Methods For the Structural Study Of Tubulin Anmentioning

confidence: 99%

The Mechanism of Tubulin Assembly into Microtubules: Insights from Structural Studies

et al. 2020

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“…Overall, it appears that the microtubule-bound conformation of Tau may delay aggregation. Cryogenic electron microscopy (cryo-EM) is bound to provide atomic structures of Tau bound to microtubules that were assembled from tubulin in different ways ( 34 ).…”

Section: Tau Isoformsmentioning

confidence: 99%

Tau Filaments and the Development of Positron Emission Tomography Tracers

et al. 2018

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A pathological pathway leading from soluble, monomeric to insoluble, filamentous Tau, is believed to underlie human Tauopathies. Cases of frontotemporal dementia are caused by dominantly inherited mutations in MAPT, the Tau gene. They show that dysfunction of Tau protein is sufficient to cause neurodegeneration and dementia. Extrapolation to the more common sporadic Tauopathies leads one to conclude that the pathological pathway is central to the development of all cases of disease, even if there are multiple reasons for Tau assembly. These findings are conceptually similar to those reported for beta-amyloid, alpha-synuclein and prion protein. Here, we provide an overview of Tau filaments and their positron emission tomography ligands.

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“…However, both SLDs and DARPins may compete with MAPs for tubulin binding. Indeed, SLDs target a tubulin surface that corresponds to the exterior of the microtubule (Gigant et al., 2000), where the binding sites of numerous MAPs are clustered (Nogales and Kellogg, 2017). Competition with DARPins has also been reported (Nawrotek et al., 2014, Sharma et al., 2016).…”

Section: Introductionmentioning

confidence: 99%

Selection and Characterization of Artificial Proteins Targeting the Tubulin α Subunit

et al. 2019

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Summary Microtubules are cytoskeletal filaments of eukaryotic cells made of αβ-tubulin heterodimers. Structural studies of non-microtubular tubulin rely mainly on molecules that prevent its self-assembly and are used as crystallization chaperones. Here we identified artificial proteins from an αRep library that are specific to α-tubulin. Turbidity experiments indicate that these αReps impede microtubule assembly in a dose-dependent manner and total internal reflection fluorescence microscopy further shows that they specifically block growth at the microtubule (−) end. Structural data indicate that they do so by targeting the α-tubulin longitudinal surface. Interestingly, in one of the complexes studied, the α subunit is in a conformation that is intermediate between the ones most commonly observed in X-ray structures of tubulin and those seen in the microtubule, emphasizing the plasticity of tubulin. These α-tubulin-specific αReps broaden the range of tools available for the mechanistic study of microtubule dynamics and its regulation.

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Challenges and opportunities in the high-resolution cryo-EM visualization of microtubules and their binding partners

Cited by 18 publications

References 34 publications

The Mechanism of Tubulin Assembly into Microtubules: Insights from Structural Studies

The Mechanism of Tubulin Assembly into Microtubules: Insights from Structural Studies

Tau Filaments and the Development of Positron Emission Tomography Tracers

Selection and Characterization of Artificial Proteins Targeting the Tubulin α Subunit

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