Hyperproduction of phosphate-binding protein, phoS, and pre-phoS proteins in Escherichia coli carrying a cloned phoS gene.
A large amount of phosphate-binding protein, the phoS gene product, accumulated in the periplasmic space of the cells when an Escherichia coli strain carrying a multicopy plasmid containing a chromosomal fragment of the phoS-phoT region (pSN507) was grown in a low-phosphate medium. When the same strain carrying a plasmid containing only the phoS gene (pSN518 or pSN5182) was grown in low-phosphate medium, phosphate-binding protein accumulated in the periplasm, and in addition a larger protein accumulated in the non-periplasmic fraction. The apparent Mr of this protein and the phosphate-binding protein were 39000 and 35000 respectively, as judged by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. This larger protein showed immunological cross-reaction with the phosphate-binding protein. The 39000-Mr protein was also detected in cells carrying pSN507 when the proteins were pulse-labeled with radioactive amino acids. From these findings, together with the fact that this protein is recovered from the membrane fraction, we conclude that this protein is an unsecreted precursor protein of the phosphate-binding protein. Kinetics and regulation of accumulation of these proteins were studied. This system will be useful for preparation and purification of the precursor protein for biochemical studies in relation to the mechanism of protein secretion.
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005798:Genes, Bacterial; D028044:Phosphate-Binding Proteins; D011498:Protein Precursors / Q000096:biosynthesis*
D001426:Bacterial Proteins; D002352:Carrier Proteins; D028044:Phosphate-Binding Proteins; D011498:Protein Precursors