Comprehensive Characterization of fucAO Operon Activation in Escherichia coli

Wildtype cells cannot grow on L-1,2-propanediol, as the operon within the fucose ( ) regulon is thought to be silent in the absence of L-fucose. Little information is available concerning the transcriptional regulation of this operon. Here, we first confirm that operon expression is highly inducible by fucose and is primarily attributable to the upstream operon promoter, while the promoter within the 3'-end of is weak and uninducible. Using 5'RACE, we identify the actual transcriptional start site (TSS) of the main operon promoter, refuting the originally proposed TSS. Several lines of evidence are provided showing that the locus is within a transcriptionally repressed region on the chromosome. Operon activation is dependent on FucR and Crp but not SrsR. Two Crp-cAMP binding sites previously found in the regulatory region are validated, where the upstream site plays a more critical role than the downstream site in operon activation. Furthermore, two FucR binding sites are identified, where the downstream site near the first Crp site is more important than the upstream site. Operon transcription relies on Crp-cAMP to a greater degree than on FucR. Our data strongly suggest that FucR mainly functions to facilitate the binding of Crp to its upstream site, which in turn activates the promoter by efficiently recruiting RNA polymerase.