Different Functionals Domain of SpastinM87V Affect Cellular Microtubule Cutting
Cell Biology
Volume 7, Issue 1, June 2019, Pages: 1-9
Received: Jan. 14, 2019; Accepted: Feb. 25, 2019; Published: Mar. 20, 2019
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Authors
Yang Yuhao, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Zhang Guowei, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Ji Zhisheng, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Cai Zhenbin, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Liu Qiuling, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Li Shaojin, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
Lin Hongsheng, Department of Orthopedics, the First Affiliated Hospital of Jinan University, Guangzhou, China
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Abstract
The research is aimed to explore the effect of different functional domains of SpastinM87V on microtubule cutting of Hela cells. First, the Spastin gene was extracted and its mutants were constructed and identified. Then, Spastin and its mutants were transfected into Hela cells and their cutting effects on microtubules were observed. The results showed that SpastinM87V and its truncated prokaryotic plasmids, including GST-SpastinN197, GST-Spastin△AAA, GST-Spastin△N1 and GST-Spastin△N2 were constructed, and the corresponding fusion proteins were expressed in vitro. In addition, SpastinM87V and its truncated eukaryotic plasmids including GFP-SpastinN197, GFP-Spastin△AAA, GFP-Spastin△N1 and GFP-Spastin△N2 were successfully constructed and introduced into Hela cells. At last, the microtubules of Hela cells could be cut into small fragments by SpastinM87V and Spastin△N1. Furthermore, the fluorescence intensity of the microtubules of Hela cells in the SpastinM87V and Spastin△N1 groups were significantly weaker than in the control group. In this way, Spastin is mainly involved in cell microtubule cutting, and the cutting function by SpastinM87V must contain complete microtubule binding- domain (MTBD) and cutting domain (AAA domain).
Keywords
SpastinM87V, Functional Domain, Protein Expression, Microtubule Cutting
To cite this article
Yang Yuhao, Zhang Guowei, Ji Zhisheng, Cai Zhenbin, Liu Qiuling, Li Shaojin, Lin Hongsheng, Different Functionals Domain of SpastinM87V Affect Cellular Microtubule Cutting, Cell Biology. Vol. 7, No. 1, 2019, pp. 1-9. doi: 10.11648/j.cb.20190701.11
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Copyright © 2019 Authors retain the copyright of this article.
This article is an open access article distributed under the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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