Understanding the yeast host cell response to recombinant membrane protein production

Abstract

Membrane proteins are drug targets for a wide range of diseases. Having access to appropriate samples for further research underpins the pharmaceutical industry's strategy for developing new drugs. This is typically achieved by synthesizing a protein of interest in host cells that can be cultured on a large scale, allowing the isolation of the pure protein in quantities much higher than those found in the protein's native source. Yeast is a popular host as it is a eukaryote with similar synthetic machinery to that of the native human source cells of many proteins of interest, while also being quick, easy and cheap to grow and process. Even in these cells, the production of human membrane proteins can be plagued by low functional yields; we wish to understand why. We have identified molecular mechanisms and culture parameters underpinning high yields and have consolidated our findings to engineer improved yeast host strains. By relieving the bottlenecks to recombinant membrane protein production in yeast, we aim to contribute to the drug discovery pipeline, while providing insight into translational processes.

A yeast expression system for functional and pharmacological studies of the malaria parasite Ca2+/H+ antiporter

“…Additionally, recent advances seeking to overcome the difficulties intrinsic to the expression of foreign proteins in yeast, particularly membrane proteins, have rendered transgenic strains with altered ribosomal content that seems to tolerate and much improve heterologous gene overexpression [56,57]. The successful expression of PfCHA in S. cerevisiae and its localization to the yeast vacuole permits a very tractable system for further functional and pharmacological studies as exemplified here in multi-well whole-cell cation transport and inhibitory assays.…”

Playing catch-up with Escherichia coli: using yeast to increase success rates in recombinant protein production experiments

“…In the last few years, significant advances have been made in this second approach by understanding how a yeast cell responds to the stress of producing a recombinant protein at a molecular level, and how this information can be used to identify improved host strains (Bonander et al, 2009; Ashe and Bill, 2011; Bawa et al, 2011; Lee et al, 2012). Since S. cerevisiae is particularly amenable to studying the mechanistic basis of high-yielding recombinant protein production experiments using the tools of systems and synthetic biology, its more routine use is an obvious way to produce less tractable proteins recombinantly (Drew et al, 2008).…”

Which Yeast Species Shall I Choose? Saccharomyces cerevisiae Versus Pichia pastoris (Review)

“…A respiratory strain of S. cerevisiae, TM6*, has been reported to have improved yield properties for both recombinant soluble and membrane proteins on account of its altered metabolism (24,33). Its improved biomass yields, which are achieved at the expense of ethanol production, result in an increased volumetric yield of recombinant protein (34). www.yeastgenome.org), which is a scientific database of yeast molecular biology and genetics.…”

“…(Insert Table 3 (40,53). For this reason, the ability to increase "per cell" yields is an area of active research, not only in P. pastoris but also in other host cells (34). P. pastoris is a methylotroph, which has two endogenous copies of the AOX gene; AOX1 expression accounts for more than 90 % of the enzyme in the cell whilst AOX2 expression constitutes less than 10 % (see Chapter 15).…”