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Biological Sciences
Research Highlights

May 2010

Strategy to Quantify, Purify Surface Proteins Also Shows Effects on Protein Translocation

Proteins expressed by Shewanella have novel roles in cell substrates

Haizhen Zhang
Haizhen Zhang
Membrane Protein Schematic strategy of membrane protein enrichment and quantification. Enlarge Image

Results. It's always good when you can get two discoveries for the price of one. A strategy developed by scientists at Pacific Northwest National Laboratory to quantify and purify proteins on the surface membranes of cells has also revealed other proteins that have potentially novel roles in cell substrates. Even more important, the researchers also found that deletion of a type II secretion protein had minimal effects on total protein expression, but significant effects on protein translocation to the cell membrane. Their results will appear in the Journal of Proteome Research.

Why it matters. Surface membrane proteins are essential for maintaining normal biological functions in cells, and often are the "first responders" to environmental stimuli. Despite their biological significance, membrane proteins can be low in abundance and insoluble, making them challenging to quantify and purify. Developing a strategy that can probe changes in membrane protein abundance will improve the understanding of overall biological cellular functions.

Methods. The PNNL team met this challenge by first enriching surface membrane proteins expressed by Shewanella oneidensis MR-1 using a membrane-impermeable chemical probe, which allowed labeling of the surface exposed peptides. By linking this method with post-digestion stable isotope labeling, the surface proteins can be quantified. The team identified about 400 proteins, of which 79% were predicted to be localized in the membrane. The successful determination of membrane protein abundance change caused by genetic deletion of one of their translocation pathways further demonstrated the specificity and sensitivity of this strategy in quantifying the membrane proteome abundance.

Acknowledgments. This work was supported by the U.S. Department of Energy Office of Biological and Environmental Research's (DOE-BER's) Genomics Science Program. Portions of this work used capabilities developed by the DOE-BER and National Center for Research Resources. Proteomics analysis was performed in the Environmental Molecular Sciences Laboratory (EMSL), a DOE-BER national scientific user facility located at PNNL. The research team included Haizhen Zhang, Mary Lipton, Roslyn Brown, Wei-Jun Qian, Matt Monroe, Sam Purvine, Ron Moore, Marina Gritsenko, Liang Shi, Margie Romine, Jim Fredrickson, Ljiljana Paša-Tolic, and Dick Smith.

Reference. Zhang H, RN Brown, W Qian, ME Monroe, SO Purvine, RJ Moore, MA Gritsenko, L Shi, MF Romine, JK Fredrickson, L Paša-Tolic, RD Smith, and MS Lipton. 2010. "Quantitative Analysis of Cell Surface Membrane Proteins using Membrane-impermeable Chemical Probe Coupled with 18O Labeling." Journal of Proteome Research 9(5):2160-2169.  DOI:10.1021/pr9009113.

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