Global Topology Analysis of the Escherichia coli Inner Membrane Proteome

The protein complement of cellular membranes is notoriously resistant to standard proteomic analysis and structural studies. As a result, membrane proteomes remain ill-defined. Here, we report a global topology analysis of the Escherichia coli inner membrane proteome. Using C-terminal tagging with the alkaline phosphatase and green fluorescent protein, we established the periplasmic or cytoplasmic locations of the C termini for 601 inner membrane proteins. By constraining a topology prediction algorithm with this data, we derived high-quality topology models for the 601 proteins, providing a firm foundation for future functional studies of this and other membrane proteomes. We also estimated the overexpression potential for 397 green fluorescent protein fusions; the results suggest that a large fraction of all inner membrane proteins can be produced in sufficient quantities for biochemical and structural work.

[1]  G. Heijne.  The distribution of positively charged residues in bacterial inner membrane proteins correlates with the trans‐membrane topology , 1986, The EMBO journal.

[2]  J. Beckwith,et al.  A genetic approach to analyzing membrane protein topology. , 1986, Science.

[3]  E. Myers,et al.  Basic local alignment search tool. , 1990, Journal of molecular biology.

[4]  L. Tisa,et al.  Membrane topology of the ArsB protein, the membrane subunit of an anion-translocating ATPase. , 1992, The Journal of biological chemistry.

[5]  I. Paulsen,et al.  The SMR family: a novel family of multidrug efflux proteins involved with the efflux of lipophilic drugs , 1996, Molecular microbiology.

[6]  G. Heijne,et al.  Divergent evolution of membrane protein topology: the Escherichia coli RnfA and RnfE homologues. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[7]  Koreaki Ito,et al.  Dislocation of membrane proteins in FtsH‐mediated proteolysis , 1999, The EMBO journal.

[8]  Gregory J. Phillips,et al.  Green Fluorescent Protein Functions as a Reporter for Protein Localization in Escherichia coli , 2000, Journal of bacteriology.

[9]  D. Fu,et al.  Structure of a glycerol-conducting channel and the basis for its selectivity. , 2000, Science.

[10]  A. Krogh,et al.  Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. , 2001, Journal of molecular biology.

[11]  A. Yamaguchi,et al.  Analysis of a Complete Library of Putative Drug Transporter Genes in Escherichia coli , 2001, Journal of bacteriology.

[12]  G. von Heijne,et al.  Green fluorescent protein as an indicator to monitor membrane protein overexpression in Escherichia coli , 2001, FEBS letters.

[13]  R. Dutzler,et al.  X-ray structure of a ClC chloride channel at 3.0 Å reveals the molecular basis of anion selectivity , 2002, Nature.

[14]  Johan Nilsson,et al.  Rapid topology mapping of Escherichia coli inner-membrane proteins by prediction and PhoA/GFP fusion analysis , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[15]  Christopher G Tate,et al.  Three‐dimensional structure of the bacterial multidrug transporter EmrE shows it is an asymmetric homodimer , 2003, The EMBO journal.

[16]  A. Krogh,et al.  Reliability measures for membrane protein topology prediction algorithms. , 2003, Journal of molecular biology.

[17]  Johan Nilsson,et al.  Experimentally based topology models for E. coli inner membrane proteins , 2004, Protein science : a publication of the Protein Society.

[18]  Bert van den Berg,et al.  X-ray structure of a protein-conducting channel , 2004, Nature.

[19]  G. Chang,et al.  Structure of the multidrug resistance efflux transporter EmrE from Escherichia coli. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[20]  Robert M. Stroud,et al.  Mechanism of Ammonia Transport by Amt/MEP/Rh: Structure of AmtB at 1.35 Å , 2004, Science.

[21]  C. Tate,et al.  New insights into the structure and oligomeric state of the bacterial multidrug transporter EmrE: an unusual asymmetric homo‐dimer , 2004, FEBS letters.

[22]  Masafumi Arai,et al.  Internal gene duplication in the evolution of prokaryotic transmembrane proteins. , 2004, Journal of molecular biology.

[23]  S. White.  The progress of membrane protein structure determination , 2004, Protein science : a publication of the Protein Society.

[24]  宁北芳,et al.  疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A , 2005 .

[25]  P. Nordlund,et al.  An efficient strategy for high‐throughput expression screening of recombinant integral membrane proteins , 2005, Protein science : a publication of the Protein Society.