title: Dimeric lac repressors exhibit phase-dependent co-operativity.
abstract: Transcription of the lac operon in Escherichia coli is repressed by the binding of Lac repressor (LacR) to lac operator O1, a pseudo-palindromic sequence centred 11 bp downstream of the transcription start. Full repression of the wild-type promoter by wild-type, tetrameric LacR requires the presence of at least two operator sequences that must not only be in close proximity to O1, 401 bp and 92 bp for the auxiliary operators O2 and O3, respectively, but must also be present on the same side of the DNA helix. LacR mutants lacking the C-terminal heptad repeat and thus only capable of dimer formation still repress, but at a much reduced level. Their repression of the lac promoter is comparable to repression by tetrameric LacR when both auxiliary operators are destroyed. We have examined the residual repression, by dimeric LacR, of a series of constructs containing a CAP-independent promoter and two lac operators, O1 and Oid, separated by a series of spacers increasing in size by single base-pair increments. Surprisingly, repression of these constructs still exhibits phase dependence. The periodicity of maxima is similar to the helical repeat of DNA in vivo, as measured by phase-dependent repression with tetrameric LacR, although the magnitude of repression is much smaller than that obtained in previous experiments with tetrameric LacR. Two additional variants of dimeric LacR with altered C termini that were tested also show phase dependence. Control experiments show that the presence of O1 is required for repression in this system. In the absence of O1, occupancy of the auxiliary operator does not lead to repression. The magnitudes of repression maxima correlate best with the overall basic nature of the C terminus. Weak, unspecific contacts by this region with DNA seem sufficient to explain the observed periodicity. It remains to be seen whether additional factors are also involved in this residual repression.
authors.affiliation: Institut für Genetik der Universität zu Köln, Köln, Weyertal 121, 50931, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Müller
authors.forename: A
authors.initials: A
authors.lastname: Barker
authors.forename: S
authors.initials: S
authors.lastname: Oehler
authors.forename: B
authors.initials: B
authors.lastname: Müller-Hill
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D007763:Lac Operon; D056968:Lac Repressors; D008969:Molecular Sequence Data; D009154:Mutation; D009875:Operator Regions, Genetic; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D012097:Repressor Proteins