"Antiparallel" DNA loop in gal repressosome visualized by atomic force microscopy

Research output: Contribution to journalJournal articleResearchpeer-review

Konstantin Virnik, Yuri L Lyubchenko, Mikhail A Karymov, Paul Dahlgren, Michael Y Tolstorukov, Szabolcs Semsey, Victor B Zhurkin, Sankar Adhya

DNA looping is often involved in positive and negative regulation of gene transcription in both prokaryotes and eukaryotes. The transcription of the gal operon of Escherichia coli from two overlapping promoters P1 and P2 is negatively regulated via Gal repressosome assembly. It involves binding of two dimeric Gal repressor proteins (GalR) to two operators, O(E) and O(I), flanking the two promoters, and formation of 113 bp DNA loop due to tetramerization of the two bound GalR dimers. The process requires negatively supercoiled DNA and the presence of the histone-like protein HU. Previous modeling of the repressosome based on evaluation of DNA elastic energy suggested a mutual antiparallel, rather than parallel, orientation of the two gal operators in an under-twisted DNA loop. To visualize the Gal loop by atomic force microscopy (AFM), plasmid DNA molecules were constructed with increased distance between the two operators. The AFM results demonstrated the formation of an antiparallel DNA loop in the Gal repressosome consistent with our earlier hypothesis. Importantly, the overall shape of the GalR mediated loop proved to be indistinguishable from that in the chimerical loop of the same size containing two lac operators (instead of two gal operators) and formed by LacI. In addition, a possibility of the gal operon repression mediated by GalR in the absence of HU was shown in the new DNA constructs. Implications of these findings for the DNA structural organization in bacterial nucleoid are discussed.

Original languageEnglish
JournalJournal of Molecular Biology
Volume334
Issue number1
Pages (from-to)53-63
Number of pages11
ISSN0022-2836
DOIs
Publication statusPublished - 14 Nov 2003
Externally publishedYes

    Research areas

  • Bacterial Proteins, DNA, Bacterial, DNA-Binding Proteins, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Macromolecular Substances, Microscopy, Atomic Force, Nucleic Acid Conformation, Operon, Promoter Regions, Genetic, Repressor Proteins, Journal Article

ID: 163917975