Abstract
We developed a protein-proximity assay in yeast based on fusing a histone lysine methyltransferase onto a bait and its substrate onto a prey. Upon binding, the prey is stably methylated and detected by methylation-specific antibodies. We applied this approach to detect varying interaction affinities among proteins in a mitogen-activated protein kinase pathway and to detect short-lived interactions between protein phosphatase 2A and its substrates that have so far escaped direct detection.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Cell Cycle Proteins / metabolism
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Histone-Lysine N-Methyltransferase / metabolism*
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Humans
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Methylation
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Mitogen-Activated Protein Kinases / metabolism
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Protein Interaction Mapping / methods*
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Protein Phosphatase 2 / metabolism
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / metabolism
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Substrate Specificity
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TOR Serine-Threonine Kinases / metabolism
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Tacrolimus Binding Proteins / metabolism
Substances
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CDC55 protein, S cerevisiae
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Cell Cycle Proteins
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Saccharomyces cerevisiae Proteins
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Histone-Lysine N-Methyltransferase
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TOR Serine-Threonine Kinases
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Mitogen-Activated Protein Kinases
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Protein Phosphatase 2
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Tacrolimus Binding Proteins