End Binding Proteins Are Obligatory Dimers
Open Access
- 6 September 2013
- journal article
- research article
- Published by Public Library of Science (PLoS) in PLOS ONE
- Vol. 8 (9), e74448
- https://doi.org/10.1371/journal.pone.0074448
Abstract
End binding (EB) proteins are responsible for the recruitment of an array of microtubule plus-end tracking proteins (+TIPs) to growing microtubules ends. EBs encompass an N-terminal calponin homology domain that confers microtubule tip tracking activity to the protein. The C-terminal domain of EBs contains a coiled coil that mediates the parallel dimerization of EB monomers. This part of the protein is also responsible for partner binding. While dimerization is not essential for microtubule tip tracking by EBs it is a prerequisite for +TIP partner binding. The concentration of EBs in cells has been estimated to be in the range of hundreds of nanomoles. In contrast, in in vitro single molecule experiments EB concentrations of subnanomoles are employed. From a mechanistic point of view it is important to assess the oligomerization state of EBs at physiologically and experimentally relevant protein concentrations, in particular if the goal of a study is to model the behavior of EB-dependent dynamic +TIP networks. Here we have determined the stability of the EB1 and EB3 dimers using multi-angle light scattering and fluorescence analytical ultracentrifugation. We show that these EBs form stable dimers and do not dissociate even at very low nanomolar concentrations. The dimers remained stable at both room temperature as well as at the physiologically relevant temperature of 37°C. Together, our results reveal that EBs are obligatory dimers, a conclusion that has implications for the mechanistic understanding of these key proteins involved in the orchestration of dynamic protein networks at growing microtubule ends.Keywords
This publication has 21 references indexed in Scilit:
- +TIPs: SxIPping along microtubule endsTrends in Cell Biology, 2012
- Plus-End-Tracking Proteins and Their Interactions at Microtubule EndsCurrent Biology, 2010
- Structural and mechanistic insights into microtubule end-binding proteinsCurrent Opinion in Cell Biology, 2010
- Molecular Insights into Mammalian End-binding Protein HeterodimerizationPublished by Elsevier BV ,2010
- An EB1-Binding Motif Acts as a Microtubule Tip Localization SignalCell, 2009
- Mammalian end binding proteins control persistent microtubule growthThe Journal of cell biology, 2009
- Tracking the ends: a dynamic protein network controls the fate of microtubule tipsNature Reviews Molecular Cell Biology, 2008
- Reconstitution of a microtubule plus-end tracking system in vitroNature, 2007
- Structural Basis of Microtubule Plus End Tracking by XMAP215, CLIP-170, and EB1Molecular Cell, 2007
- Structural insights into the EB1?APC interactionThe EMBO Journal, 2004