Davey John, Ladds Graham
Department of Clinical Sciences, University of Warwick, Coventry, UK.
Methods Mol Biol. 2011;746:113-31. doi: 10.1007/978-1-61779-126-0_7.
In this chapter, we describe methods to heterologously express G protein-coupled receptors (GPCRs) in the fission yeast Schizosaccharomyces (Sz.) pombe. GPCRs regulate a diverse range of biological processes in all eukaryotic cells, including plants, insects, humans, and yeast. The high degree of conservation between GPCRs from different organisms has facilitated the development of a large number of model systems to enable study of this pharmaceutically important family of cell-surface receptors. Of the many model systems available for investigating GPCRs, yeast have proven to be one of the more attractive. Yeasts' amenability to both genetic and biochemical manipulation, a reduced number of endogenous GPCRs and their relative low culturing costs has facilitated their use in many high-throughput drug screens. Given the high number of detailed methods relating to the expression of GPCRs within budding yeast, we have focused our attention on the use of fission yeast as a model system. We describe the methods used and provide examples from our own experiences of expressing a number of human GPCRs in Sz. pombe cells.
在本章中，我们描述了在裂殖酵母粟酒裂殖酵母中异源表达G蛋白偶联受体（GPCR）的方法。GPCR在所有真核细胞中调节多种生物过程，包括植物、昆虫、人类和酵母。来自不同生物体的GPCR之间的高度保守性促进了大量模型系统的开发，以便能够研究这个在药学上具有重要意义的细胞表面受体家族。在众多可用于研究GPCR的模型系统中，酵母已被证明是更具吸引力的系统之一。酵母易于进行遗传和生化操作，内源性GPCR数量减少且培养成本相对较低，这促进了它们在许多高通量药物筛选中的应用。鉴于与芽殖酵母中GPCR表达相关的详细方法数量众多，我们将注意力集中在使用裂殖酵母作为模型系统上。我们描述了所使用的方法，并提供了我们自己在粟酒裂殖酵母细胞中表达多种人类GPCR的经验示例。
