《MPEC_yeast_screen》.pdf

doi:10.1016/j.jmb.2008.11.021 J. Mol. Biol. (2009) 385, 820–830 Available online at Selecting Optimum Eukaryotic Integral Membrane Proteins for Structure Determination by Rapid Expression and Solubilization Screening Min Li1†, Franklin A. Hays2 ⁎†, Zygy Roe-Zurz1 , Linda Vuong1 , Libusha Kelly3,4,5,6,7 , Chi-Min Ho1 , Renée M. Robbins1 , Ursula Pieper3 , Joseph D. OConnell III 2 , Larry J. W. Miercke1,2, Kathleen M. Giacomini3,5,6,7 , Andrej Sali3,5,6,7 and Robert M. Stroud1,2,3⁎ 1Membrane Protein Expression A medium-throughput approach is used to rapidly identify membrane Center, University of California proteins from a eukaryotic organism that are most amenable to expression at San Francisco, San Francisco, in amounts and quality adequate to support structure determination. The CA 94158-2517, USA goal was to expand knowledge of new membrane protein structures based 2 on proteome-wide coverage. In the first phase, membrane proteins from the Department of Biochemistry budding yeast Saccharomyces cerevisiae were selected for homologous and Biophysics, University of expression in S. cerevisiae, a system that can be adapted to expression of California at San Francisco, membrane proteins from other eukaryotes. We performed medium-scale San Francisco, CA 94158-2517, USA expression and solubilization tests on 351 rationally selected membrane proteins from S. cerevisiae. These targets are inclusive of all annotated and 3Center for the Structure of unannotated membrane protein families within the organisms membrane Membrane Proteins, University proteome. Two hundred seventy-two tar