Yeast as a tool for membrane protein production and structure determination

• Although the majority of eukaryotic MEMBRANE PROTEIN structures are DERIVED FROM PROTEINS produced in HEK293 and insect cells, the authors show here the importance of yeast as a production host and its role as an essential player in the production of eukaryotic membrane proteins for structural and functional analysis. Membrane proteins are challenging targets to functionally and structurally characterize. An enduring bottleneck in their study is the reliable production of sufficient yields of stable protein. Here, we evaluate all eukaryotic membrane protein production experiments that have supported the deposition of a high-resolution structure. We focused on the most common yeast host systems, Saccharomyces cerevisiae and Pichia pastoris. The first high-resolution structure of a membrane protein produced in yeast was described in 1999 and today there are 186 structures of alpha-helical membrane proteins, representing 101 unique proteins from 37 families. Homologous and heterologous production are equally common in S. cerevisiae, while heterologous production dominates in P. pastoris, especially of human proteins, which represent about one-third of the total. Investigating protein engineering approaches (78 proteins from seven families) demonstrated that the majority contained a polyhistidine tag for purification, typically at the C-terminus of the protein. Codon optimization and truncation of hydrophilic extensions were also common approaches to improve yields. We conclude that yeast remains a useful production host for the study of alpha-helical membrane proteins.

Ämnesord

NATURVETENSKAP  -- Biologi -- Mikrobiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Microbiology (hsv//eng)

TEKNIK OCH TEKNOLOGIER  -- Industriell bioteknik -- Annan industriell bioteknik (hsv//swe)

ENGINEERING AND TECHNOLOGY  -- Industrial Biotechnology -- Other Industrial Biotechnology (hsv//eng)

Nyckelord

membrane protein production

Saccharomyces cerevisiae

Pichia pastoris

x-ray-structure

cytochrome bc(1) complex

monoamine-oxidase-a

cryo-em

structure

crystal-structure

saccharomyces-cerevisiae

k+ channel

pichia-pastoris

functional-analysis

atomic-structure

Biotechnology & Applied Microbiology

Microbiology

Mycology

membrane protein production

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