XMAP215 polymerase activity is built by combining multiple tubulin-binding TOG domains and a basic lattice-binding region
Open Access
- 31 January 2011
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 108 (7), 2741-2746
- https://doi.org/10.1073/pnas.1016498108
Abstract
XMAP215/Dis1 family proteins positively regulate microtubule growth. Repeats at their N termini, called TOG domains, are important for this function. While TOG domains directly bind tubulin dimers, it is unclear how this interaction translates to polymerase activity. Understanding the functional roles of TOG domains is further complicated by the fact that the number of these domains present in the proteins of different species varies. Here, we take advantage of a recent crystal structure of the third TOG domain from Caenorhabditis elegans, Zyg9, and mutate key residues in each TOG domain of XMAP215 that are predicted to be important for interaction with the tubulin heterodimer. We determined the contributions of the individual TOG domains to microtubule growth. We show that the TOG domains are absolutely required to bind free tubulin and that the domains differentially contribute to XMAP215’s overall affinity for free tubulin. The mutants’ overall affinity for free tubulin correlates well with polymerase activity. Furthermore, we demonstrate that an additional basic region is important for targeting to the microtubule lattice and is critical for XMAP215 to function at physiological concentrations. Using this information, we have engineered a “bonsai” protein, with two TOG domains and a basic region, that has almost full polymerase activity.This publication has 33 references indexed in Scilit:
- Review of the mechanism of processive actin filament elongation by forminsCell Motility, 2009
- Implications for Kinetochore-Microtubule Attachment from the Structure of an Engineered Ndc80 ComplexCell, 2008
- XMAP215 Is a Processive Microtubule PolymeraseCell, 2008
- Structural Basis of Microtubule Plus End Tracking by XMAP215, CLIP-170, and EB1Molecular Cell, 2007
- Mechanism and Function of Formins in the Control of Actin AssemblyAnnual Review of Biochemistry, 2007
- Crystal Structure of a TOG Domain: Conserved Features of XMAP215/Dis1-Family TOG Domains and Implications for Tubulin BindingStructure, 2007
- Assembly dynamics of microtubules at molecular resolutionNature, 2006
- Stu2p binds tubulin and undergoes an open-to-closed conformational changeThe Journal of cell biology, 2006
- The Interaction of TOGp with Microtubules and TubulinJournal of Biological Chemistry, 2000
- Rate constants for the reactions of ATP- and ADP-actin with the ends of actin filaments.The Journal of cell biology, 1986