Strategies for the cloning and expression of membrane proteins.
Despite the determination of thousands of high-resolution structures of soluble proteins, many features of integral membrane proteins render them difficult targets for the structural biologist. Among these, the most important challenge is in expressing sufficient quantities of active protein to support downstream purification and structure determination efforts. Over 190 unique membrane protein structures have now been solved, and noticeable trends in successful expression strategies are beginning to emerge. A number of groups have also explored high-throughput (HTP) methods for membrane protein expression, with varying degrees of success. Here we review the current state of expressing membrane proteins for functional and structural studies. We first survey successful methods that have already yielded levels of membrane protein expression sufficient for structure determination. HTP methods are also examined since these aim to explore large numbers of targets and can predict reasonable starting points for many membrane proteins. Since HTP techniques may fail, particularly for certain classes of eukaryotic targets, detailed strategies for the expression of two prominent classes of eukaryotic protein families, G-protein-coupled receptors and ion channels, are also summarized.
Koth
Christopher M M
CM
Department of Structural Biology, Genentech, South San Francisco, California 94080, USA.
Payandeh
Jian
J
D003001:Cloning, Molecular; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*
D008565:Membrane Proteins