4.6 Article

Membrane Association of the PTEN Tumor Suppressor: Molecular Details of the Protein-Membrane Complex from SPR Binding Studies and Neutron Reflection

Journal

PLOS ONE
Volume 7, Issue 4, Pages -

Publisher

PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0032591

Keywords

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Funding

  1. National Institutes of Health [1R01 NS021716, 1P01 AG032131]
  2. National Science Foundation [CHE 0724082, 1216827]
  3. Direct For Mathematical & Physical Scien
  4. Division Of Chemistry [1216827] Funding Source: National Science Foundation

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The structure and function of the PTEN phosphatase is investigated by studying its membrane affinity and localization on in-plane fluid, thermally disordered synthetic membrane models. The membrane association of the protein depends strongly on membrane composition, where phosphatidylserine (PS) and phosphatidylinositol diphosphate (PI(4,5)P-2) act pronouncedly synergistic in pulling the enzyme to the membrane surface. The equilibrium dissociation constants for the binding of wild type (wt) PTEN to PS and PI(4,5)P-2 were determined to be K-d similar to 12 mu M and 0.4 mu M, respectively, and K-d similar to 50 nM if both lipids are present. Membrane affinities depend critically on membrane fluidity, which suggests multiple binding sites on the protein for PI(4,5)P-2. The PTEN mutations C124S and H93R show binding affinities that deviate strongly from those measured for the wt protein. Both mutants bind PS more strongly than wt PTEN. While C124S PTEN has at least the same affinity to PI(4,5)P-2 and an increased apparent affinity to PI(3,4,5) P-3, due to its lack of catalytic activity, H93R PTEN shows a decreased affinity to PI(4,5)P-2 and no synergy in its binding with PS and PI(4,5)P-2. Neutron reflection measurements show that the PTEN phosphatase scoots'' along the membrane surface (penetration <5 angstrom) but binds the membrane tightly with its two major domains, the C2 and phosphatase domains, as suggested by the crystal structure. The regulatory C-terminal tail is most likely displaced from the membrane and organized on the far side of the protein similar to 60 angstrom away from the bilayer surface, in a rather compact structure. The combination of binding studies and neutron reflection allows us to distinguish between PTEN mutant proteins and ultimately may identify the structural features required for membrane binding and activation of PTEN.

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