Ratiocinative screen of eukaryotic integral membrane protein expression and solubilization for structure determination

Franklin A. Hays; Zygy Roe-Zurz; Min Li; Libusha Kelly; Franz Gruswitz; Andrej Sali; Robert M. Stroud

doi:10.1007/s10969-008-9046-7

Ratiocinative screen of eukaryotic integral membrane protein expression and solubilization for structure determination

Franklin A. Hays, Zygy Roe-Zurz, Min Li, Libusha Kelly, Franz Gruswitz, Andrej Sali, Robert M. Stroud

Research output: Contribution to journal › Review article › peer-review

9 Scopus citations

Abstract

Persistent hurdles impede the successful determination of high-resolution crystal structures of eukaryotic integral membrane proteins (IMP). We designed a high-throughput structural genomics oriented pipeline that seeks to minimize effort in uncovering high-quality, responsive non-redundant targets for crystallization. This "discovery-oriented" pipeline sidesteps two significant bottlenecks in the IMP structure determination pipeline: expression and membrane extraction with detergent. In addition, proteins that enter the pipeline are then rapidly vetted by their presence in the included volume on a size-exclusion column-a hallmark of well-behaved IMP targets. A screen of 384 rationally selected eukaryotic IMPs in baker's yeast Saccharomyces cerevisiae is outlined to demonstrate the results expected when applying this discovery-oriented pipeline to whole-organism membrane proteomes.

Original language	English (US)
Pages (from-to)	9-16
Number of pages	8
Journal	Journal of Structural and Functional Genomics
Volume	10
Issue number	1
DOIs	https://doi.org/10.1007/s10969-008-9046-7
State	Published - Mar 2009
Externally published	Yes

Keywords

Discovery-oriented screen
Eukaryotic integral membrane protein
Membrane protein structure
Saccharomyces cerevisiae
Structural genomics

ASJC Scopus subject areas

Structural Biology
Biochemistry
Genetics

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10.1007/s10969-008-9046-7

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title = "Ratiocinative screen of eukaryotic integral membrane protein expression and solubilization for structure determination",

abstract = "Persistent hurdles impede the successful determination of high-resolution crystal structures of eukaryotic integral membrane proteins (IMP). We designed a high-throughput structural genomics oriented pipeline that seeks to minimize effort in uncovering high-quality, responsive non-redundant targets for crystallization. This {"}discovery-oriented{"} pipeline sidesteps two significant bottlenecks in the IMP structure determination pipeline: expression and membrane extraction with detergent. In addition, proteins that enter the pipeline are then rapidly vetted by their presence in the included volume on a size-exclusion column-a hallmark of well-behaved IMP targets. A screen of 384 rationally selected eukaryotic IMPs in baker's yeast Saccharomyces cerevisiae is outlined to demonstrate the results expected when applying this discovery-oriented pipeline to whole-organism membrane proteomes.",

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note = "Funding Information: Acknowledgements This review describes, in part, a broad screen which was published previously in The Journal of Molecular Biology [30]—we are grateful to all co-authors involved in bringing that project to fruition. This work was supported by the N.I.H. Roadmap Center Grant P50 GM073210 (to R.M.S.), Specialized Center for the Protein Structure Initiative Grant U54 GM074929 (to R.M.S. and A.S.), National Research Service Award F32 GM078754 (to F.A.H.) and a Sandler Biomedical Research postdoctoral fellowship (to F.A.H.).",

year = "2009",

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doi = "10.1007/s10969-008-9046-7",

language = "English (US)",

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AU - Roe-Zurz, Zygy

AU - Li, Min

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AU - Gruswitz, Franz

AU - Sali, Andrej

AU - Stroud, Robert M.

N1 - Funding Information: Acknowledgements This review describes, in part, a broad screen which was published previously in The Journal of Molecular Biology [30]—we are grateful to all co-authors involved in bringing that project to fruition. This work was supported by the N.I.H. Roadmap Center Grant P50 GM073210 (to R.M.S.), Specialized Center for the Protein Structure Initiative Grant U54 GM074929 (to R.M.S. and A.S.), National Research Service Award F32 GM078754 (to F.A.H.) and a Sandler Biomedical Research postdoctoral fellowship (to F.A.H.).

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Ratiocinative screen of eukaryotic integral membrane protein expression and solubilization for structure determination

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