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Spastin's Microtubule-Binding Properties and Comparison to Katanin
Spastin and katanin are ring-shaped hexameric AAA ATPases that sever microtubules, and thus crucially depend on a physical interaction with microtubules. For the first time, we report here the microtubule binding properties of spastin at the single-molecule level, and compare them to katanin. Micros...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3521757/ https://www.ncbi.nlm.nih.gov/pubmed/23272056 http://dx.doi.org/10.1371/journal.pone.0050161 |
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author | Eckert, Thomas Le, Doan Tuong-Van Link, Susanne Friedmann, Lena Woehlke, Günther |
author_facet | Eckert, Thomas Le, Doan Tuong-Van Link, Susanne Friedmann, Lena Woehlke, Günther |
author_sort | Eckert, Thomas |
collection | PubMed |
description | Spastin and katanin are ring-shaped hexameric AAA ATPases that sever microtubules, and thus crucially depend on a physical interaction with microtubules. For the first time, we report here the microtubule binding properties of spastin at the single-molecule level, and compare them to katanin. Microscopic fluorescence assays showed that human spastin bound to microtubules by ionic interactions, and diffused along microtubules with a diffusion coefficient comparable to katanin. The microscopic measurement of landing and dissociation rates demonstrated the ionic character of the interaction, which could be mapped to a patch of three lysine residues outside of the catalytic domain of human spastin. This motif is not conserved in Drosophila spastin or katanin, which also bound by non-catalytic parts of the protein. The binding affinities of spastin and katanin were nucleotide-sensitive, with the lowest affinities under ADP,, the highest under ATP-γS conditions. These changes correlated with the formation of higher oligomeric states, as shown in biochemical experiments and electron microscopic images. Vice versa, the artificial dimerization of human spastin by addition of a coiled coil led to a constitutively active enzyme. These observations suggest that dimer formation is a crucial step in the formation of the active complex, and thus the severing process by spastin. |
format | Online Article Text |
id | pubmed-3521757 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-35217572012-12-27 Spastin's Microtubule-Binding Properties and Comparison to Katanin Eckert, Thomas Le, Doan Tuong-Van Link, Susanne Friedmann, Lena Woehlke, Günther PLoS One Research Article Spastin and katanin are ring-shaped hexameric AAA ATPases that sever microtubules, and thus crucially depend on a physical interaction with microtubules. For the first time, we report here the microtubule binding properties of spastin at the single-molecule level, and compare them to katanin. Microscopic fluorescence assays showed that human spastin bound to microtubules by ionic interactions, and diffused along microtubules with a diffusion coefficient comparable to katanin. The microscopic measurement of landing and dissociation rates demonstrated the ionic character of the interaction, which could be mapped to a patch of three lysine residues outside of the catalytic domain of human spastin. This motif is not conserved in Drosophila spastin or katanin, which also bound by non-catalytic parts of the protein. The binding affinities of spastin and katanin were nucleotide-sensitive, with the lowest affinities under ADP,, the highest under ATP-γS conditions. These changes correlated with the formation of higher oligomeric states, as shown in biochemical experiments and electron microscopic images. Vice versa, the artificial dimerization of human spastin by addition of a coiled coil led to a constitutively active enzyme. These observations suggest that dimer formation is a crucial step in the formation of the active complex, and thus the severing process by spastin. Public Library of Science 2012-12-13 /pmc/articles/PMC3521757/ /pubmed/23272056 http://dx.doi.org/10.1371/journal.pone.0050161 Text en © 2012 Eckert et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Eckert, Thomas Le, Doan Tuong-Van Link, Susanne Friedmann, Lena Woehlke, Günther Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title | Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title_full | Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title_fullStr | Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title_full_unstemmed | Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title_short | Spastin's Microtubule-Binding Properties and Comparison to Katanin |
title_sort | spastin's microtubule-binding properties and comparison to katanin |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3521757/ https://www.ncbi.nlm.nih.gov/pubmed/23272056 http://dx.doi.org/10.1371/journal.pone.0050161 |
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