Comparison of protein binding to DNA in vivo and in vitro: defining an effective intracellular target.
We have quantitatively evaluated the affinity of a set of target sites for the integration host factor (IHF) protein of Escherichia coli by their performance as competitors in an electrophoretic mobility shift assay. We also determined how well each of these sites is filled by IHF in vivo. The data show that several natural sites have an affinity not much greater than that required for intracellular occupancy. The data also indicate that very little of the IHF in a cell is present as free protein available for binding, suggesting that binding to non-specific targets dominates the operation of this system. The correlation between in vitro affinity and in vivo occupancy provides a ready means to assess the likely physiological significance of putative IHF sites. It also provides a general method to assess the importance of non-specific interactions by DNA binding proteins inside a cell.
Yang
S W
SW
Laboratory of Molecular Biology, NIMH, Bethesda MD 20892-4034, USA.
Nash
H A
HA
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D014644:Genetic Variation; D034982:Integration Host Factors; D007700:Kinetics; D008969:Molecular Sequence Data; D011485:Protein Binding
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D034982:Integration Host Factors