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Operator‐bound GalR dimers close DNA loops by direct interaction: tetramerization and inducer binding
Author(s) -
Semsey Szabolcs,
Geanacopoulos Mark,
Lewis Dale E.A.,
Adhya Sankar
Publication year - 2002
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/cdf431
Subject(s) - biology , inducer , dna , operator (biology) , plasma protein binding , biophysics , repressor , binding site , dna binding protein , biochemistry , microbiology and biotechnology , transcription factor , gene
The assembly of the Gal repressosome, a higher order nucleoprotein complex that represses transcription of the gal operon in Escherichia coli , involves the formation of a DNA loop encompassing the promoter segment. GalR dimers bound to two spatially separated operators, O E and O I , specifically interact with the histone‐like protein HU and close the loop in supercoiled DNA. We isolated and characterized a GalR mutant containing an amino acid substitution (R282L) that can repress transcription in the absence of HU and supercoiled DNA both in vivo and in vitro . Repression involves the same DNA looping; deletion of either O E or O I makes the mutant GalR ineffective in repression. This and other results suggest that the R282L substitution increases the normal affinity between two DNA‐bound GalR dimers, allowing looping. We conclude that GalR dimers interact directly and do not use HU as an adaptor in loop closure; HU and DNA supercoiling act in concert to stabilize the GalR tetramer. The stronger GalR–GalR interaction also made the gal transcription non‐inducible, suggesting that the inducer binding acts by modulating tetramerization.