In Vitro Repression of the gal Promoters by GalR and HU Depends on the Proper Helical Phasing of the Two Operators

• Published in: The Journal of biological chemistry Vol. 277; no. 4; pp. 2498 - 2504
• Main Authors: Lewis, Dale E.A., Adhya, Sankar
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 25.01.2002; American Society for Biochemistry and Molecular Biology
• Subjects: AHU protein; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Base Sequence; Binding Sites; DNA - metabolism; DNA looping; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - metabolism; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Escherichia coli - metabolism; Escherichia coli Proteins; GalR protein; HU protein; Models, Genetic; Molecular Sequence Data; Mutation; Plasmids - metabolism; Promoter Regions, Genetic; Protein Binding; Repressor Proteins - genetics; Repressor Proteins - metabolism; Time Factors; Transcription, Genetic
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M108456200

Repression of transcription initiation from the two gal promoters, P1 and P2, requires binding of GalR protein to two flanking operators,OE and OI, binding of HU to a site, hbs, located between the two operators, and supercoiled DNA template. Previous experiments suggested that repression involves the interaction of two DNA-bound GalR proteins, which generates a 113-bp DNA loop encompassing the promoter region. Interaction between two DNA-bound proteins would be allowed if the binding sites on DNA are properly aligned. To test the idea that the observed repression of gal transcription in vitro is mediated by DNA looping, we investigated the effect of changing the relative angular orientation of OEand OI in the DNA helix. We found that repression is a periodic function of the distance between the two operator sites. Since repression recurred commensurate with DNA helical repeat, we conclude that the observed in vitro repression is mediated by DNA looping and the in vitro conditions reflect the in vivo situation.