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.