Stable Expression of Multisubunit Protein Complexes in Mammalian Cells
There are advantages and disadvantages to each of the different protein expression systems available. Whereas ease, speed, and expense are important factors in determining which type of expression system to use, the complexity of the protein to be expressed and the kinds of questions addressed will often be the factors that ultimately determine which system is used. Our interest has been in expressing the nicotinic acetylcholine receptor (AChR). This protein is a heterologous multisubunit complex composed of four different subunits in the stoichiometry α2βγδ. In addition to the complex subunit composition, each subunit undergoes several posttranslational modifications and spans the membrane a multiple number of times. In order to reconstitute all of the functional properties of the AChR, the four subunits must be present and at least some of the posttranslational modifications must be executed correctly. Thus, whichever gene transfer technique one uses to express AChRs, the ability to introduce all four of the subunits into the same cell and to introduce them into a cell type that is capable of performing the necessary modifications, is of considerable importance. A mechanical technique such as microinjection would clearly be the easiest solution to the problem of multiple subunits.
KeywordsLong Terminal Repeat Sodium Butyrate Selectable Marker Gene AChR Cluster Muscle Cell Line
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