Influence of cyclic AMP, agmatine, and a novel protein encoded by a flanking gene on speB (agmatine ureohydrolase) in Escherichia coli.

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RESUMO

The speB gene of Escherichia coli encodes agmatine ureohydrolase (AUH), a putrescine biosynthetic enzyme. The speB gene is transcribed either from its own promoter or as a polycistronic message from the promoter of the speA gene encoding arginine decarboxylase. Two open reading frames (ORF1 and ORF2) are present on the strand complementary to speB; approximately 90% of ORF2 overlaps the speB coding region. Analysis of transcriptional and translational fusions of ORF1 or ORF2 to lacZ revealed that ORF1 encoded a novel protein while ORF2 was not transcribed. Deletion of ORF1 from a plasmid containing ORF1, ORF2, and speB reduced the activity of AUH by 83%. In contrast, the presence of plasmid-encoded ORF1 caused an 86% increase in chromosomally encoded AUH activity. ORF1 did not stimulate alkaline phosphatase expressed from a phi(speB-phoA) transcriptional fusion encoded on the same plasmid. Western analysis (immunoblot) of a phi(ORF1-lacZ) translational fusion revealed that ORF1 encodes a 25.3-kDa protein. Agmatine induced transcription of phi(speB-phoA) but not phi(speA-phoA) fusions. Consequently, agmatine affects selection between the monocistronic and the polycistronic modes of speB transcription. In contrast, cyclic AMP (cAMP) repressed AUH activity of chromosomally encoded AUH but had no effect on plasmid-borne speB nor phi(speB-phoA). It is concluded that ORF1 encodes a protein which is a posttranscriptional regulator of speB, agmatine induces speB independent of speA, and cAMP regulates speB indirectly.

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