Single molecule study of non-specific binding kinetics of LacI in mammalian cells.

Item request has been placed! ×
Item request cannot be made. ×
loading   Processing Request
Read More Add to Saved list
  • Additional Information
    • Source:
      Publisher: Faraday Division, Royal Society of Chemistry Country of Publication: England NLM ID: 9212301 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1364-5498 (Electronic) Linking ISSN: 13596640 NLM ISO Abbreviation: Faraday Discuss Subsets: MEDLINE
    • Publication Information:
      Original Publication: London : Faraday Division, Royal Society of Chemistry, c1992-
    • Subject Terms:
      Cells/*metabolism ; DNA/*chemistry ; Lac Repressors/*chemistry; Binding Sites ; Cell Line, Tumor ; DNA/metabolism ; Humans ; Kinetics ; Lac Repressors/metabolism ; Molecular Dynamics Simulation ; Spectrometry, Fluorescence
    • Abstract:
      Many key cellular processes are controlled by the association of DNA-binding proteins (DBPs) to specific sites. The kinetics of the search process leading to the binding of DBPs to their target locus are largely determined by transient interactions with non-cognate DNA. Using single-molecule microscopy, we studied the dynamics and non-specific binding to DNA of the Lac repressor (LacI) in the environment of mammalian nuclei. We measured the distribution of the LacI-DNA binding times at non-cognate sites and determined the mean residence time to be τ(1D) = 182 ms. This non-specific interaction time, measured in the context of an exogenous system such as that of human U2OS cells, is remarkably different compared to that reported for the LacI in its native environment in E. coli (<5 ms). Such a striking difference (more than 30 fold) suggests that the genome, its organization, and the nuclear environment of mammalian cells play important roles on the dynamics of DBPs and their non-specific DNA interactions. Furthermore, we found that the distribution of off-target binding times follows a power law, similar to what was reported for TetR in U2OS cells. We argue that a possible molecular origin of such a power law distribution of residence times is the large variability of non-cognate sequences found in the mammalian nucleus by the diffusing DBPs.
    • Accession Number:
      0 (Lac Repressors)
      9007-49-2 (DNA)
    • Publication Date:
      Date Created: 20150922 Date Completed: 20160707 Latest Revision: 20151215
    • Publication Date:
      20221213
    • Accession Number:
      10.1039/c5fd00112a
    • Accession Number:
      26387491