De novo design of defined helical bundles in membrane environments

Başar Bilgiçer* and
Krishna Kumar*,†,‡

^*Department of Chemistry, Tufts University, Medford, MA 02155; and ^†Cancer Center, Tufts–New England Medical Center, Boston, MA 02110

Edited by William F. DeGrado, University of Pennsylvania, Philadelphia, PA (received for review May 11, 2004)

Abstract

Control of structure and function in membrane proteins remains a formidable challenge. We report here a new design paradigm for the self-assembly of protein components in the context of nonpolar environments of biological membranes. An incrementally staged assembly process relying on the unique properties of fluorinated amino acids was used to drive transmembrane helix–helix interactions. In the first step, hydrophobic peptides partitioned into micellar lipids. Subsequent phase separation of simultaneously hydrophobic and lipophobic fluorinated helical surfaces fueled spontaneous self-assembly of higher order oligomers. The creation of these ordered transmembrane protein ensembles is supported by gel electrophoresis, circular dichroism spectroscopy, equilibrium analytical ultracentrifugation, and fluorescence resonance energy transfer.

Footnotes

↵ ‡ To whom correspondence should be addressed. E-mail: krishna.kumar{at}tufts.edu.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: TAMRA, 5-(and-6)-carboxytetramethylrhodamine; FRET, fluorescence resonance energy transfer; NBD, 7-nitrobenz-2-oxa-1,3-diazole.
↵ § Cybotactic region has been defined as that part of a solution in the vicinity of a solute molecule in which the ordering of the solvent molecules is modified by the presence of the solute molecule.
↵ ¶ Labeled peptides were utilized for sedimentation experiments because the large extinction coefficient of the NBD chromophore allowed greater sensitivity in detection.
↵ ∥ Expressed in units of peptide/detergent mole fraction (MF).