title: Overexpression, purification, and functional analysis of recombinant human tubulin dimer.
abstract: Microtubules consisting of tubulin dimers play essential roles in various cellular functions. Investigating the structure-function relationship of tubulin dimers requires a method to prepare sufficient quantities of recombinant tubulin. To this end, we simultaneously expressed human α1- and β3-tubulin using a baculovirus-insect cell expression system that enabled the purification of 5mg recombinant tubulin per litre of cell culture. The purified recombinant human tubulin could be polymerized into microtubules that glide on a kinesin-coated glass surface. The method provides a powerful tool for in vitro functional analyses of microtubules.
authors.affiliation: Laboratory for Molecular Biophysics, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
authors.forename: Itsushi
authors.initials: I
authors.lastname: Minoura
authors.forename: You
authors.initials: Y
authors.lastname: Hachikubo
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Yamakita
authors.forename: Hiroko
authors.initials: H
authors.lastname: Takazaki
authors.forename: Rie
authors.initials: R
authors.lastname: Ayukawa
authors.forename: Seiichi
authors.initials: S
authors.lastname: Uchimura
authors.forename: Etsuko
authors.initials: E
authors.lastname: Muto
mesh_terms: D002478:Cells, Cultured; D006801:Humans; D016547:Kinesins / Q000378:metabolism; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D014404:Tubulin / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014404:Tubulin; D016547:Kinesins
keywords: Baculovirus-insect cell expression; Kinesin motility; MAP; PTM; Tubulin assembly; WT; microtubule-associated protein; posttranslational modification; wild-type