Spontaneous Reconstitution of Bovine Rhodopsin into Artificial Membranes

Abstract

Rhodopsin is a canonical G-protein-coupled receptor (GPCR) that is responsible for vision in dim light. It has the potential to serve as a high-fidelity, light-sensing molecular switch for a broad range of nanotechnologies. Previous studies revealed that the photoactivity of rhodopsin depends critically on the native lipid bilayer environment surrounding this membrane protein [1-3]. It is not yet clear how artificial membranes in synthetic systems would affect the activity of rhodopsin [4], and recent study suggests that membrane moduli may play important roles [4, 5]. Partially this uncertainty is due to the fact that it is experimentally challenging to prepare rhodopsin-supporting artificial proteomembranes with systematically varied membrane chemistry and physical properties. Here we show that bovine rhodopsin can be spontaneously reconstituted into a series of well-defined artificial membranes, including both lipid-based (i.e., liposome) and polymer-based (i.e., polymersome) membranes, via a charge-interaction-directed reconstitution (CIDR) mechanism [5, 6]. Using synchrotron small-angle X-ray scattering (SAXS), we show that n-dodecyl-β-D-maltoside solubilized bovine rhodopsin is reconstituted spontaneously to form 2-D proteomembrane arrays, which in turn are coupled along the trans-membrane direction to form a 3-D multilamellar structure. The lamellar periodicity is ∼5.7 nm, which matches closely the transmembrane dimension of rhodopsin. Using time-resolved UV-visible spectroscopy, we are currently examining the photoactivity of embedded bovine rhodopsin, and its dependency on the surface charge states and membrane moduli of the artificial membrane. [1] M. F. Brown (1997) Curr. Top. Membr. 44, 285-356. [2] A.V. Botelho et al. (2006) Biophys. J. 91, 4464-4477. [3] A.V. Struts et al. (2014) Meth. Mol. Biol. (in press). [4] V. Subramaniam et al. (2005) JACS 127, 5320-5321. [5] L.J. Kuang et al. (2014) ACS Nano 8, 537-545. [6] D.B. Hua et al. (2011) JACS 133, 2354-2357.