Abstract
This chapter describes a general method for expressing several G-protein α subunits in Escherichia coli (E. coli) at levels 10-100 times higher than achieved previously. G proteins are a family of guanine nucleotide-binding regulatory proteins that link a large number of cell surface receptors to regulation of several intracellular effectors. The chapter describes a method for purification of the recombinant proteins by affinity chromatography on a resin containing chelated Ni2+ after addition of an amino-terminal hexahistidine tag to the recombinant protein. Such purification is rapid and results in the isolation of highly purified protein in a single step. Furthermore, the introduction of a tobacco etch virus (TEV) polyprotein cleavage site between the hexahistidine tag and the G-protein α subunit permits the efficient removal of the tag by recombinant TEV protease.
Original language | English (US) |
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Pages (from-to) | 146-164 |
Number of pages | 19 |
Journal | Methods in Enzymology |
Volume | 237 |
Issue number | C |
DOIs | |
State | Published - Jan 1 1994 |
ASJC Scopus subject areas
- Biochemistry
- Molecular Biology