Cell-free translation of integral membrane proteins into unilamelar liposomes

Technology

Cloning

Summary

Wheat germ cell-free translation is shown to be an effective method to produce integral membrane proteins in the presence of unilamelar liposomes.

Description

In this chapter, we describe the expression vectors, preparation of mRNA, two types of cell-free translation reactions performed in the presence of liposomes, a simple and highly efficient purification of intact proteoliposomes using density gradient ultracentrifugation, and some of the types of characterization studies that are facilitated by this facile preparative approach. The in vitro transfer of newly translated, membrane proteins into liposomes compatible with direct measurements of their catalytic function is contrasted with existing approaches to extract membrane proteins from biological membranes using detergents and subsequently transfer them back to liposomes for functional studies.

Publication

Goren MA, Nozawa A, Makino S, Wrobel RL, Fox BG. Cell-free translation of integral membrane proteins into unilamelar liposomes. Methods Enzymol 2009;463:647-73. PubMed ID: 19892197 | Search SBKB Publications portal

Contact

Brian G. Fox (bgfox@biochem.wisc.edu)

Transmembrane Protein Center

Availability

Email for details

Link




share on Facebook Tweet This share on LinkedIn add Google Bookmark add to delicious Share on reddit share on StumbleUpon Digg this


Last edited:Mon 10 Mar 2014 - 3 years, 3 months ago