Abstract
We attempted the selection of a site for DNA binding to a transcription factor, PhaR, from Paracoccus denitrificans expressed by cell-free protein synthesis, from a random oligonucleotide library on microbeads that was constructed by the emulsion PCR technique. PhaR is a repressor protein in P. denitrificans that binds to the phaP promoter region. We acquired three types of PhaR-binding DNA fragment using this system. The selected fragments contained a perfect PhaR binding consensus site (TGC I), a sequence similar to that of TGC I, and another PhaR binding site (TGC II).
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Bacterial Proteins / genetics*
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Bacterial Proteins / metabolism*
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Binding Sites
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Cell-Free System
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DNA-Binding Proteins / genetics*
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DNA-Binding Proteins / metabolism*
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Flow Cytometry / methods*
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Microspheres
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Paracoccus denitrificans / genetics*
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Paracoccus denitrificans / metabolism*
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Peptide Library
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Promoter Regions, Genetic / genetics
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Protein Engineering / methods*
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Repressor Proteins / genetics*
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Repressor Proteins / metabolism*
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Transcription Factors / genetics
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Transcription Factors / metabolism*
Substances
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Bacterial Proteins
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DNA-Binding Proteins
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Peptide Library
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Repressor Proteins
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Transcription Factors