Transcriptional regulation of molybdoenzyme synthesis in Escherichia coli in response to molybdenum: ModE-molybdate, a repressor of the modABCD (molybdate transport) operon is a secondary transcriptional activator for the hyc and nar operons

Escherichia coli growing under anaerobic conditions produces several molybd oenzymes, such as formate hydrogenlyase (formate to H-2 and CO2; hyc and fd hF genes) and nitrate reductase (narGHJI genes). Synthesis of these molybdo entymes, even in the presence of the cognate transcriptional activators and effecters, requires molybdate in the medium. Besides the need for molybdop terin cofactor synthesis, molybdate is also required for transcription of t he genes encoding these molybdoenzymes. In E. coli, ModE was previously ide ntified as a repressor controlling transcription of the operon encoding mol ybdate transport components (modABCD). In this work, the ModE protein was a lso found to be a required component in the activation of hyc-lacZ to an op timum level, but only in the presence of molybdate. Mutant ModE proteins wh ich are molybdate-independent for repression of modA-lacZ also restored hyc -lacZ expression to the wild-type level even in the absence of molybdate. N itrate-dependent enhancement of transcription of narX-lacZ was completely a bolished in a modE mutant. Nitrate-response by narG-lacZ and narK-lacZ was reduced by about 50% in a modE mutant. DNase I footprinting experiments rev ealed that the ModE protein binds the hyc promoter DNA in the presence of m olybdate, ModE-molybdate also protected DNA in the intergenic region betwee n narXL and narK from DNase I hydrolysis. DNA sequences (5' TAYAT 3' and 5' GTTA 3') found in ModE-molybdate-protected modABCD operator DNA were also found in the ModE-molybdate-protected region of hyc promoter DNA (5' GTTA-7 bp-CATAT 3') and narX-narK intergenic region (5' GTTA-7 bp-TACAT 3'). Base d on these results, a working model is proposed in which ModE-molybdate ser ves as a secondary transcriptional activator of both the hyc and narXL oper ons which are activated primarily by the transcriptional activators, FhIA a nd NarL, respectively.