Fluorinated and hemifluorinated surfactants as alternatives to detergents for membrane protein cell-free synthesis.
Hemifluorinated and fluorinated surfactants are lipophobic and, as such, non-detergent. Although they do not solubilize biological membranes, they can, after conventional solubilization, substitute for detergents to keep membrane proteins soluble, which generally improves their stability [Breyton, Chabaud, Chaudier, Pucci and Popot (2004) FEBS Lett. 564, 312-318]. In the present study, we show that (hemi)fluorinated surfactants can be used for in vitro synthesis of membrane proteins: they do not interfere with protein synthesis, and they provide a suitable environment for MscL, a pentameric mechanosensitive channel, to fold and oligomerize to its native functional state. Following synthesis, both types of surfactants can be used to deliver MscL directly to pre-formed lipid vesicles. The electrophysiological activity of MscL synthesized in vitro in the presence of either hemi- or per-fluorinated surfactant is similar to that of the protein expressed in vivo.
Park
Kyu-Ho
KH
Laboratoire de Physico-chimie Moléculaire des Membranes Biologiques, UMR 7099, CNRS and Université Paris-7, Institut de Biologie Physico-Chimique, CNRS FRC 550, F-75005 Paris, France.
Berrier
Catherine
C
Lebaupain
Florence
F
Pucci
Bernard
B
Popot
Jean-Luc
JL
Ghazi
Alexandre
A
Zito
Francesca
F
D002462:Cell Membrane / Q000502:physiology; D002474:Cell-Free System; D003902:Detergents; D004926:Escherichia coli / Q000502:physiology; D006845:Hydrocarbons, Fluorinated; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000502:physiology; D018408:Patch-Clamp Techniques; D011510:Proteolipids / Q000737:chemistry; D013501:Surface-Active Agents / Q000737:chemistry*
D003902:Detergents; D006845:Hydrocarbons, Fluorinated; D008565:Membrane Proteins; D011510:Proteolipids; D013501:Surface-Active Agents; C020485:proteoliposomes