Abstract
The reorganization of microtubules in mitosis, meiosis, and development requires the microtubule-severing activity of katanin. Katanin is a heterodimer composed of an ATPase associated with diverse cellular activities (AAA) subunit and a regulatory subunit. Microtubule severing requires ATP hydrolysis by katanin's conserved AAA ATPase domains. Whereas other AAA ATPases form stable hexamers, we show that katanin forms only a monomer or dimers of heterodimers in solution. Katanin oligomers consistent with hexamers of heterodimers or heterododecamers were only observed for an ATP hydrolysis-deficient mutant in the presence of ATP. X-ray structures of katanin's AAA ATPase in monomeric nucleotide-free and pseudo-oligomeric ADP-bound states revealed conformational changes in the AAA subdomains that explained the structural basis for the instability of the katanin heterododecamer. We propose that the rapid dissociation of katanin AAA oligomers may lead to an autoinhibited state that prevents inappropriate microtubule severing or that cyclical disassembly into heterodimers may critically contribute to the microtubule-severing mechanism.
Highlights
The reorganization of microtubules in mitosis, meiosis, and development requires the microtubule-severing activity of katanin
Our size-exclusion chromatography (SEC)-MALS results indicate that WT KATNAL1/B1-con80 does not assemble beyond dimers of heterodimers at concentrations up to 50 M
Our SEC-MALS analysis clearly demonstrated that human KATNAL1/B1-con80 does not assemble beyond a dimer of heterodimers, even at concentrations up to 50 M
Summary
The reorganization of microtubules in mitosis, meiosis, and development requires the microtubule-severing activity of katanin. 2 The abbreviations used are: MT, microtubule; MIT, microtubule-interacting and trafficking; VPS4, vacuolar protein sorting–associated protein 4; con, conserved p80; NSF, N-ethylmaleimide–sensitive fusion; SEC, sizeexclusion chromatography; SEC-MALS, SEC-coupled multiangle light scattering; AMPPNP, 5Ј-adenylyl-,␥-imidodiphosphate; SAD, single-wavelength anomalous dispersion; NBD, nucleotide-binding domain; HBD, 4-helix bundle domain; N-Hlx, N-terminal helix; C-Hlx, C-terminal helix; RMSD, root-mean-square deviation; TEV, tobacco etch virus; GST, glutathione S-transferase; PDB, Protein Data Bank.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
