CONTROL OF POLYPHOSPHATE METABOLISM IN GENETICALLY-ENGINEERED ESCHERICHIA-COLI

The polyphosphate operon of Escherichia coli was overexpressed on a hi gh copy plasmid under control of its native promoter. This operon cont ains the genes coding far polyphosphate kinase (PPK) and polyphosphata se (PPX), the enzymes responsible for polyphosphate synthesis and degr adation, respectively The plasmid was engineered for improved stabilit y by addition of the parB locus. The polyphosphate content of cells co ntaining this plasmid was significantly higher than that of wildtype E . coli. Cells were subjected to phosphate limitation following a perio d of exponential growth, and then shifted back to high-phosphate condi tions. Polyphosphate reserves were utilized during phosphate starvatio n and partially replenished after the shift PPK activity increased dur ing phosphate starvation and dropped after the shift while PPX activit y was highest when phosphate was in surplus. A ppk::lacZ reporter gene construct was used to investigate transcription from the ppk promoter . Although expression doubled upon phosphate starvation, it is unlikel y that the Pho regulon is involved since both phoB and phoU mutant hos t strains gave similar responses. (C) 1998 Elsevier Science Inc.