The structural and functional consequences of the interaction of the vinculin tail domain with F-actin and PIP2

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  • March 19, 2019
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  • Thompson, Peter
    • Affiliation: School of Medicine, Department of Biochemistry and Biophysics
Abstract
  • Vinculin is an essential, highly-conserved eukaryotic scaffolding protein. It localizes to focal adhesions and adherens juctions, where it assists in physically linking the actin cytoskeleton to the adhesive structure. Loss of vinculin causes embryonic lethality with observable cardiovascular and neural defects. Vinculin is comprised of three domains: a large helical head domain, an unstructured proline-rich linker, and a helical tail. Vinculin functions as a scaffold, alternating between an autoinhibited conformation in which it cannot bind ligands, and an open conformation in which it is free to bind ligands at all three domains. The tail domain binds a variety of ligands, two of which are F-actin and phosphatidylinositol 4,5-bisphosphate (PIP2). Both interactions lack good structural models, which has hindered the development of tools to understand the specific biological functions these interactions play. Using a variety of biochemical and biophysical techniques and tools, we have developed new structural models for these interactions. The PIP2 model shows how the basic residues in helices 1 and 2 specifically recognize PIP2, while basic residues on helix 3 are responsible for membrane association. Actin-binding, conversely, is driven by a hydrophobic patch on helix 4. The application of cryo-electron microscopy has also revealed some of the conformational changes that take place in the tail domain when it binds F-actin: helix 4 straightens and helix 1 is discharged from the helix bundle. Additionally, we characterize the backbone dynamics of Vt by nuclear magnetic resonance. These experiments reveal the presence of μs-ms motions that reside where the C-terminal arm interacts with the helix bundle. These motions may be involved in the conformational change that takes place upon actin binding.
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  • In Copyright
Advisor
  • Collins, Edward
  • Errede, Beverly
  • Cox, Adrienne
  • Lee, Andrew
  • Campbell, Sharon
Degree
  • Doctor of Philosophy
Degree granting institution
  • University of North Carolina at Chapel Hill Graduate School
Graduation year
  • 2015
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  • Chapel Hill, NC
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  • There are no restrictions to this item.
Date uploaded
  • June 23, 2015

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