Highly efficient selenomethionine labeling of recombinant proteins produced in mammalian cells Journal Article
| Authors: | Barton, W. A.; Tzvetkova Robev, D.; Erdjument-Bromage, H.; Tempst, P.; Nikolov, D. B. |
| Article Title: | Highly efficient selenomethionine labeling of recombinant proteins produced in mammalian cells |
| Abstract: | The advent of the multiwavelength anomalous diffraction phasing method has significantly accelerated crystal structure determination and has become the norm in protein crystallography. This method allows researchers to take advantage of the anomalous signal fromdiverse atoms, but the dominant method for derivative preparation is selenomethionine substitution. Several generally applicable, high-efficiency labeling protocols have been developed for use in the bacterial, yeast, and baculovirus/insect cell expression systems but not for mammalian tissue culture. As a large number of proteins of biomedical importance can only be produced in yields sufficient for X-ray diffraction experiments in mammalian expression systems, it becomes all the more important to develop such protocols. We therefore evaluated several variables that play roles in determining incorporation levels and report here a simple protocol for selenomethionine modification of proteins in mammalian cells routinely yielding >90% labeling efficiency. Published by Cold Spring Harbor Laboratory Press. Copyright © 2006 The Protein Society. |
| Keywords: | controlled study; human cell; mammalia; amino acid substitution; cell line; bacteria (microorganisms); genetic transfection; cell culture; recombinant proteins; recombinant protein; cell culture techniques; crystal structure; crystallography, x-ray; crystallization; matrix assisted laser desorption ionization time of flight mass spectrometry; receptor, tie-2; culture media; staining and labeling; chemical modification; mammal cell; spectrometry, mass, matrix-assisted laser desorption-ionization; x ray diffraction; crystallography; selenomethionine; chemical labeling; mammalian cell culture; multiple wavelength anomalous diffraction; protein crystallography; protein labeling |
| Journal Title: | Protein Science |
| Volume: | 15 |
| Issue: | 8 |
| ISSN: | 0961-8368 |
| Publisher: | Wiley Blackwell |
| Date Published: | 2006-08-01 |
| Start Page: | 2008 |
| End Page: | 2013 |
| Language: | English |
| DOI: | 10.1110/ps.062244206 |
| PUBMED: | 16823037 |
| PROVIDER: | scopus |
| PMCID: | PMC2242577 |
| DOI/URL: | |
| Notes: | --- - "Cited By (since 1996): 9" - "Export Date: 4 June 2012" - "CODEN: PRCIE" - "Source: Scopus" |
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