Abstract
Using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunological techniques, we have compared the synthesis of the phoA protein (alkaline phosphatase) and the phoS protein (phosphate-binding protein) in response to the level of phosphate in the medium in different genetic backgrounds containing the known alkaline phosphatase control mutations. Both proteins are produced in excess phosphate media in a phoR1a- strain, whereas neither protein is produced in a phoB- strain even under derepression conditions. In four different phoR1c- strains, however, the phoA product cannot be detected in extracts of cells obtained from any growth condition, whereas the phoS product is produced in both excess and limiting phosphate media. It is not yet known if phoR1c- mutants are a special class of mutations within the phoB gene or whether they occur in a separate cistron involved in alkaline phosphatase regulation. From these results we conclude that the expression of the phoA gene is not always co-regulated with expression of the phoS gene product. We have determined that the phoS protein is a component of periplasmic protein band P4 described by Morris et al. (1974). The phoS product lacks sulfur-containing amino acids and is extractable by treatment with polymyxin sulfate. The other component of band P4 contains methionine and/or cysteine and is not extracted by polymyxin sulfate treatment. Like the phoS and phoA proteins, its synthesis is sensitive to the concentration of phosphate in the growth medium. In addition, the existence of a new class of periplasmic proteins synthesized at maximum rate in high phosphate media is demonstrated.
Full text
PDF![595](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/aac0feee404a/jbacter00314-0609.png)
![596](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/35006df711cd/jbacter00314-0610.png)
![597](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/b53d2545c327/jbacter00314-0611.png)
![598](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/d5ee0382a7ae/jbacter00314-0612.png)
![599](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/42e237e5bcfb/jbacter00314-0613.png)
![600](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/2d716e5545f3/jbacter00314-0614.png)
![601](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/fbf3e3f06f64/jbacter00314-0615.png)
![602](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/e6d0da3d4fca/jbacter00314-0616.png)
![603](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/599f3045071e/jbacter00314-0617.png)
![604](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/2b7151ea9d37/jbacter00314-0618.png)
![605](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/7efd020bcb05/jbacter00314-0619.png)
![606](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/4afdbc271470/jbacter00314-0620.png)
![607](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/6cbf95d0fb50/jbacter00314-0621.png)
![608](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/d529694db359/jbacter00314-0622.png)
![609](https://cdn.ncbi.nlm.nih.gov/pmc/blobs/535a/233093/40cd48302723/jbacter00314-0623.png)
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Aono H., Otsuji N. Genetic mapping of regulator gene phoS for alkaline phosphatase in Escherichia coli. J Bacteriol. 1968 Mar;95(3):1182–1183. doi: 10.1128/jb.95.3.1182-1183.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bennett R. L., Malamy M. H. Arsenate resistant mutants of Escherichia coli and phosphate transport. Biochem Biophys Res Commun. 1970 Jul 27;40(2):496–503. doi: 10.1016/0006-291x(70)91036-3. [DOI] [PubMed] [Google Scholar]
- Bracha M., Yagil E. A ne type of alkaline phosphatase-negative mutants in Escherichia coli K12. Mol Gen Genet. 1973 Mar 27;122(1):53–60. doi: 10.1007/BF00337973. [DOI] [PubMed] [Google Scholar]
- Bracha M., Yagil E. Genetic mapping of the phoR regulator gene of alkaline phosphatase in Escherichia coli. J Gen Microbiol. 1969 Nov;59(1):77–81. doi: 10.1099/00221287-59-1-77. [DOI] [PubMed] [Google Scholar]
- Brickman E., Beckwith J. Analysis of the regulation of Escherichia coli alkaline phosphatase synthesis using deletions and phi80 transducing phages. J Mol Biol. 1975 Aug 5;96(2):307–316. doi: 10.1016/0022-2836(75)90350-2. [DOI] [PubMed] [Google Scholar]
- Cerny G., Teuber M. Differential release of periplasmic versus cytoplasmic enzymes from Escherichia coli B by polymixin B. Arch Mikrobiol. 1971;78(2):166–179. doi: 10.1007/BF00424873. [DOI] [PubMed] [Google Scholar]
- Demerec M., Adelberg E. A., Clark A. J., Hartman P. E. A proposal for a uniform nomenclature in bacterial genetics. Genetics. 1966 Jul;54(1):61–76. doi: 10.1093/genetics/54.1.61. [DOI] [PMC free article] [PubMed] [Google Scholar]
- ECHOLS H., GAREN A., GAREN S., TORRIANI A. Genetic control of repression of alkaline phosphatase in E. coli. J Mol Biol. 1961 Aug;3:425–438. doi: 10.1016/s0022-2836(61)80055-7. [DOI] [PubMed] [Google Scholar]
- GAREN A., ECHOLS H. Genetic control of induction of alkaline phosphatase synthesis in E. coli. Proc Natl Acad Sci U S A. 1962 Aug;48:1398–1402. doi: 10.1073/pnas.48.8.1398. [DOI] [PMC free article] [PubMed] [Google Scholar]
- GAREN A., OTSUJI N. ISOLATION OF A PROTEIN SPECIFIED BY A REGULATOR GENE. J Mol Biol. 1964 Jun;8:841–852. doi: 10.1016/s0022-2836(64)80165-0. [DOI] [PubMed] [Google Scholar]
- Gerdes R. G., Rosenberg H. The relationship between the phosphate-binding protein and a regulator gene product from Escherichia coli. Biochim Biophys Acta. 1974 May 10;351(1):77–86. doi: 10.1016/0005-2795(74)90066-x. [DOI] [PubMed] [Google Scholar]
- HORIUCHI T., HORIUCHI S., MIZUNO D. A possible negative feedback phenomenon controlling formation of alkaline phosphomonoesterase in Escherichia coli. Nature. 1959 May 30;183(4674):1529–1530. doi: 10.1038/1831529b0. [DOI] [PubMed] [Google Scholar]
- Kida S. The biological function of the R2a regulatory gene for alkaline phosphatase in Escherichia coli. Arch Biochem Biophys. 1974 Jul;163(1):231–237. doi: 10.1016/0003-9861(74)90473-1. [DOI] [PubMed] [Google Scholar]
- Kreuzer K., Pratt C., Torriani A. Genetic analysis of regulatory mutants of alkaline phosphatase of E. coli. Genetics. 1975 Nov;81(3):459–468. doi: 10.1093/genetics/81.3.459. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LIN E. C., KOCH J. P., CHUSED T. M., JORGENSEN S. E. Utilization of L-alpha-glycerophosphate by Escherichia coli without hydrolysis. Proc Natl Acad Sci U S A. 1962 Dec 15;48:2145–2150. doi: 10.1073/pnas.48.12.2145. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
- MALAMY M. H., HORECKER B. L. RELEASE OF ALKALINE PHOSPHATASE FROM CELLS OF ESCHERICHIA COLI UPON LYSOZYME SPHEROPLAST FORMATION. Biochemistry. 1964 Dec;3:1889–1893. doi: 10.1021/bi00900a017. [DOI] [PubMed] [Google Scholar]
- Morris H., Schlesinger M. J., Bracha M., Yagil E. Pleiotropic effects of mutations involved in the regulation of Escherichia coli K-12 alkaline phosphatase. J Bacteriol. 1974 Aug;119(2):583–592. doi: 10.1128/jb.119.2.583-592.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nossal N. G., Heppel L. A. The release of enzymes by osmotic shock from Escherichia coli in exponential phase. J Biol Chem. 1966 Jul 10;241(13):3055–3062. [PubMed] [Google Scholar]
- Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
- TORRIANI A. Influence of inorganic phosphate in the formation of phosphatases by Escherichia coli. Biochim Biophys Acta. 1960 Mar 11;38:460–469. doi: 10.1016/0006-3002(60)91281-6. [DOI] [PubMed] [Google Scholar]
- Taylor A. L., Trotter C. D. Linkage map of Escherichia coli strain K-12. Bacteriol Rev. 1972 Dec;36(4):504–524. doi: 10.1128/br.36.4.504-524.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
- Wilkins A. S. Physiological factors in the regulation of alkaline phosphatase synthesis in Escherichia coli. J Bacteriol. 1972 May;110(2):616–623. doi: 10.1128/jb.110.2.616-623.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Willsky G. R., Bennett R. L., Malamy M. H. Inorganic phosphate transport in Escherichia coli: involvement of two genes which play a role in alkaline phosphatase regulation. J Bacteriol. 1973 Feb;113(2):529–539. doi: 10.1128/jb.113.2.529-539.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Willsky G. R., Malamy M. H. The loss of the phoS periplasmic protein leads to a change in the specificity of a constitutive inorganic phosphate transport system in Escherichia coli. Biochem Biophys Res Commun. 1974 Sep 9;60(1):226–233. doi: 10.1016/0006-291x(74)90195-8. [DOI] [PubMed] [Google Scholar]