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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
3
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pubmed:dateCreated |
2005-5-3
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pubmed:abstractText |
We report a method of photo-cross-linking proteins in mammalian cells, which is based on site-specific incorporation of a photoreactive amino acid, p-benzoyl-L-phenylalanine (pBpa), through the use of an expanded genetic code. To analyze the cell signaling interactions involving the adaptor protein Grb2, pBpa was incorporated in its Src homology 2 (SH2) domain. The human GRB2 gene with an amber codon was introduced into Chinese hamster ovary (CHO) cells, together with the genes for the Bacillus stearothermophilus suppressor tRNA(Tyr) and a pBpa-specific variant of Escherichia coli tyrosyl-tRNA synthetase (TyrRS). The Grb2 variant with pBpa in the amber position was synthesized when pBpa was included in the growth medium. Upon exposure of cells to 365-nm light, protein variants containing pBpa in the positions proximal to the ligand-binding pocket were cross-linked with the transiently expressed epidermal growth factor (EGF) receptor in the presence of an EGF stimulus. Cross-linked complexes with endogenous proteins were also detected. In vivo photo-cross-linking with pBpa incorporated in proteins will be useful for studying protein-protein interactions in mammalian cells.
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pubmed:language |
eng
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pubmed:journal |
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pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Adaptor Proteins, Signal Transducing,
http://linkedlifedata.com/resource/pubmed/chemical/Amino Acids,
http://linkedlifedata.com/resource/pubmed/chemical/Cross-Linking Reagents,
http://linkedlifedata.com/resource/pubmed/chemical/GRB2 Adaptor Protein,
http://linkedlifedata.com/resource/pubmed/chemical/GRB2 protein, human,
http://linkedlifedata.com/resource/pubmed/chemical/Penicillin-Binding Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Receptor, Epidermal Growth Factor,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Fusion Proteins
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pubmed:status |
MEDLINE
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pubmed:month |
Mar
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pubmed:issn |
1548-7091
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pubmed:author |
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pubmed:issnType |
Print
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pubmed:volume |
2
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
201-6
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pubmed:dateRevised |
2009-11-19
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pubmed:meshHeading |
pubmed-meshheading:15782189-Adaptor Proteins, Signal Transducing,
pubmed-meshheading:15782189-Amino Acid Substitution,
pubmed-meshheading:15782189-Amino Acids,
pubmed-meshheading:15782189-Animals,
pubmed-meshheading:15782189-Bacillus,
pubmed-meshheading:15782189-CHO Cells,
pubmed-meshheading:15782189-Cricetinae,
pubmed-meshheading:15782189-Cricetulus,
pubmed-meshheading:15782189-Cross-Linking Reagents,
pubmed-meshheading:15782189-Escherichia coli,
pubmed-meshheading:15782189-GRB2 Adaptor Protein,
pubmed-meshheading:15782189-Humans,
pubmed-meshheading:15782189-Light,
pubmed-meshheading:15782189-Molecular Probe Techniques,
pubmed-meshheading:15782189-Mutagenesis, Site-Directed,
pubmed-meshheading:15782189-Penicillin-Binding Proteins,
pubmed-meshheading:15782189-Photochemistry,
pubmed-meshheading:15782189-Protein Engineering,
pubmed-meshheading:15782189-Protein Interaction Mapping,
pubmed-meshheading:15782189-Receptor, Epidermal Growth Factor,
pubmed-meshheading:15782189-Recombinant Fusion Proteins,
pubmed-meshheading:15782189-Spectrum Analysis
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pubmed:year |
2005
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pubmed:articleTitle |
Protein photo-cross-linking in mammalian cells by site-specific incorporation of a photoreactive amino acid.
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pubmed:affiliation |
Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
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pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
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