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Biophysical Journal 85:3848-3857 (2003)
© 2003 The Biophysical Society

Molecular Orientation and Conformation of Phosphatidylinositides in Membrane Mimetics Using Variable Angle Sample Spinning (VASS) NMR

Anita I. Kishore and James H. Prestegard

Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602

Correspondence: Address reprint requests to James H. Prestegard, E-mail: jpresteg{at}ccrc.uga.edu.

For many biological molecules, determining their geometry as they exist in a membrane environment is a crucial step in understanding their function. Variable angle sample spinning (VASS) NMR provides a new route to obtaining geometry information on membrane-associating molecules; it has been used here to scale and separate anisotropic contributions to phosphorus chemical shifts in NMR spectra of phosphatidylinositol phosphates. The procedure allows spectral assignment via correlation with isotropic chemical shifts and determination of a family of probable headgroup orientations via interpretation of anisotropic shift contributions. The molecules studied include phosphtidylinositol-4-phosphate (PI(4)P) and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). A membrane-like environment is provided by a dispersion of alkyl-poly(ethylene) glycols and n-alcohols that forms a field-orienting liquid crystal with a director that can be manipulated by varying the sample spinning axis. The experiments presented indicate that the variable angle sample spinning method will provide a direct approach for assignment and extraction of structural information from membrane-associating biomolecules labeled with a wider variety of NMR active isotopes.




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