K. Hardoyo,"yamada et al., PRODUCTION AND RELEASE OF POLYPHOSPHATE BY A GENETICALLY-ENGINEERED STRAIN OF ESCHERICHIA-COLI, Applied and environmental microbiology, 60(10), 1994, pp. 3485-3490
A recombinant strain of Escherichia coli MV1184, which contains plasmi
d-borne genes encoding the phosphate-specific transport (Pst) system a
nd polyphosphate (polyP) kinase, accumulated high levels of P-i, and r
eleased polyP into the medium. PolyP could be separated from the cultu
re supernatant by DEAE-Toyopearl 650M chromatography and identified by
high-resolution P-31 nuclear magnetic resonance spectroscopy. Once E.
coli recombinants accumulated high levels of polyP, they released pol
yP concomitantly with P-i uptake. PolyP release did not accompany the
decrease in the cell density, indicating that it is not simply a resul
t of cell lysis. PolyP release ceased when P-i became depleted in the
medium and resumed upon addition of P-i to the medium. When P-i uptake
was inhibited by 0.1 mM carbonyl cyanide m-chlorophenylhydrazone (CCC
P), no polyP release was observed. Furthermore, neither P-i uptake nor
polyp release occurred when cells were incubated at 4 degrees C. Thes
e findings suggest that the occurrence of polyP release is a possible
mechanism that limits a further increase in the cellular polyP concent
ration in E. coli recombinants. High-resolution P-31 nuclear magnetic
resonance spectroscopy also detected a surface pool of polyp in intact
cells of the E. coli recombinant. The polyp resonance increased when
cells were treated with EDTA and broadened upon the addition of a shif
t reagent, praseodymium. Although the mechanism of surface polyP accum
ulation is unclear, surface polyP seems to serve as the source for pol
yP release.