The chemistry of phospholipid binding by the Saccharomyces cerevisiae phosphatidylinositol transfer protein Sec14p as determined by EPR spectroscopy. Academic Article uri icon

abstract

  • The major yeast phosphatidylinositol/phosphatidylcholine transfer protein Sec14p is the founding member of a large eukaryotic protein superfamily. Functional analyses indicate Sec14p integrates phospholipid metabolism with the membrane trafficking activity of yeast Golgi membranes. In this regard, the ability of Sec14p to rapidly exchange bound phospholipid with phospholipid monomers that reside in stable membrane bilayers is considered to be important for Sec14p function in cells. How Sec14p-like proteins bind phospholipids remains unclear. Herein, we describe the application of EPR spectroscopy to probe the local dynamics and the electrostatic microenvironment of phosphatidylcholine (PtdCho) bound by Sec14p in a soluble protein-PtdCho complex. We demonstrate that PtdCho movement within the Sec14p binding pocket is both anisotropic and highly restricted and that the C5 region of the sn-2 acyl chain of bound PtdCho is highly shielded from solvent, whereas the distal region of that same acyl chain is more accessible. Finally, high field EPR reports on a heterogeneous polarity profile experienced by a phospholipid bound to Sec14p. Taken together, the data suggest a headgroup-out orientation of Sec14p-bound PtdCho. The data further suggest that the Sec14p phospholipid binding pocket provides a polarity gradient that we propose is a primary thermodynamic factor that powers the ability of Sec14p to abstract a phospholipid from a membrane bilayer.

published proceedings

  • J Biol Chem

author list (cited authors)

  • Smirnova, T. I., Chadwick, T. G., MacArthur, R., Poluektov, O., Song, L., Ryan, M. M., Schaaf, G., & Bankaitis, V. A

citation count

  • 17

complete list of authors

  • Smirnova, Tatyana I||Chadwick, Thomas G||MacArthur, Ryan||Poluektov, Oleg||Song, Likai||Ryan, Margaret M||Schaaf, Gabriel||Bankaitis, Vytas A

publication date

  • September 2006