1887

Abstract

We investigated whether the concentration of H or metal ions such as Ca, or both, on the external side of the cytoplasmic membrane is involved in coupling of folding and secretion of levansucrase by studying the modulation of each isolated event. at 30°C, in the absence of Ca, the equilibrium between the unfolded and the folded states of levansucrase was rapidly and totally displaced toward the folded state by a small pH shift from 7·4 to 6·0. Ca (> 5 mM) acted as a catalyst of folding at pH ≥ 7. pulse-chase experiments at 30°C showed that, in the absence of Ca, the rate and the yield of the second step of levansucrase secretion were strongly dependent on the external pH. In acidic growth medium (pH 5·8), secretion was efficient. In contrast, at pH ≥ 7, the presence of Ca was essential for secretion. In bacteria grown at high temperature (48°C), both external acidic pH and Ca were required for efficient secretion. Moreover, a levansucrase variant altered in its calcium affinity was efficiently secreted only under acidic growth conditions. Depending on the culture conditions, the differences in H or Ca concentrations which are maintained between the opposite sides of the energized cytoplasmic membrane could be adequate to catalyse conformational transition which could play a critical role in the second step of the protein release. These environmental parameters could also affect the yield of secretion of some other secretory proteins, leading to the hypothesis that several different secretion mechanisms could coexist in .

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1995-04-01
2024-03-29
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References

  1. Archibald A. R. 1993; The Bacillus cell envelope.. In Bacillus pp. 217–254 Edited by Harwood C. R. New York: Plenum Press;
    [Google Scholar]
  2. Baker J., Craig E. A. 1994; Heat shock proteins as molecular chaperones.. Eur J Biochem 219:11–23
    [Google Scholar]
  3. Beveridge T. J., Murray R. G. E. 1976; Uptake and retention of metals by cell walls of Bacillus subtilis. . J Bacteriol 127:1502–1518
    [Google Scholar]
  4. Beveridge T. J., Murray R. G. E. 1980; Sites of metal deposition in the cell wall of Bacillus subtilis. . J Bacteriol 141:1876–1887
    [Google Scholar]
  5. Chambert R., Petit-Glatron M. F. 1984; Hyperproduction of exocellular levansucrase by Bacillus subtilis: examination of the phenotype of a sacUh strain.. J Gen Microbiol 130:3143–3152
    [Google Scholar]
  6. Chambert R., Petit-Glatron M. F. 1988; Secretion mechanism of Bacillus subtilis levansucrase: characterization of the second step.. J Gen Microbiol 134:1205–1214
    [Google Scholar]
  7. Chambert R., Petit-Glatron M. F. 1990; Reversible thermal unfolding of Bacillus subtilis levansucrase is modulated by Fe3+ and Ca2+.. FEBS Lett 275:61–64
    [Google Scholar]
  8. Chambert R., Benyahia F., Petit-Glatron M. F. 1990; Secretion of Bacillus subtilis levansucrase. Fe(III) could act as cofactor in an efficient coupling of the folding and translocation processes.. Biochem J 265:375–382
    [Google Scholar]
  9. Dedonder R. 1966; Levansucrase from Bacillus subtilis. . Methods Enzymol 8:500–506
    [Google Scholar]
  10. Eilers M., Schatz G. 1988; Protein unfolding and the energetics of protein translocation across biological membranes.. Cell 52:481–483
    [Google Scholar]
  11. Fujishige A., Smith K. R., Silen J. L., Agard D. A. 1992; Correct folding of α-lytic protease is required for its extracellular secretion from Escherichia coli. . J Cell Biol 118:33–42
    [Google Scholar]
  12. Hemilä H., Pokkinen M., Palva I. 1992; Improving the production of E. coli β-lactamase in Bacillus subtilis: the effect of glucose, pH and temperature on the production level.. J Biotechnol 26:245–256
    [Google Scholar]
  13. Kemper M. A., Urrutia T. J., Beveridge A. L., Doyle R. J. 1993; Proton motive force may regulate cell wall associated enzymes of Bacillus subtilis. . J Bacteriol 175:5690–5696
    [Google Scholar]
  14. Khan S., Macnab R. M. 1980; Proton chemical potential, proton electrical potential and bacterial motility. . J Mol Biol 138:599–614
    [Google Scholar]
  15. Koch A. L. 1986; The pH in the neighborhood of membranes generating a protonmotive force.. J Theor Biol 120:73–84
    [Google Scholar]
  16. Kontinen V. P., Saris P., Sarvas M. 1991; A gene (prsA) of Bacillus subtilis involved in a novel late stage of protein export.. Mol Microbiol 5:1273–1283
    [Google Scholar]
  17. Kumamoto C. A. 1991; Molecular chaperones and protein translocation across the Escherichia coli inner membrane.. Mol Microbiol 5:19–22
    [Google Scholar]
  18. Langer T., Neupert W. 1991; Heat shock proteins hsp60 and hsp70: their roles in folding, assembly and membrane translocation of proteins.. Curr Top Microbiol Immunol 167:3–30
    [Google Scholar]
  19. London E. 1992; How bacterial toxins enter cells: the role of partial unfolding in membrane translocation.. Mol Microbiol 6:3277–3282
    [Google Scholar]
  20. Mäntsälä P., Zalkin H. 1979; Membrane-bound and soluble extracellular α-amylase from Bacillus subtilis. . J Biol Chem 254:8540–8547
    [Google Scholar]
  21. Maruyama K., Mikawa T., Ebashi S. 1984; Detection of calcium binding proteins by 45Ca autoradiography on nitrocellulose membranes after sodium dodecyl sulfate gel electrophoresis.. J Biochem 95:511–519
    [Google Scholar]
  22. McLaughlin S. 1977; Electrostatic potentials at membrane—solution interfaces.. Curr Top Membr Transp 9:71–144
    [Google Scholar]
  23. Minnikin D. E., Abdeolrahimzadeh H. 1974; Effect of pH on the proportions of polar lipids in chemostat cultures of Bacillus subtilis. . J Bacterio1 120:999–1003
    [Google Scholar]
  24. Otha Y., Hojo H., Aimoto S., Kobayashi T., Zhu X., Jordan F., Inouye M. 1991; Pro-peptide as an intramolecular chaperone: renaturation of denatured subtilisin E with a synthetic pro-peptide.. Mol Microbiol 5:1507–1510
    [Google Scholar]
  25. Palmen R., Driessen A. J., Hellingwerf K. J. 1994; Bioenergetic aspects of the translocation of macromolecules across bacterial membranes.. Biochim Biophys Acta 1183:417–451
    [Google Scholar]
  26. Petit-Glatron M. F., Benyahia F., Chambert R. 1987; Secretion of Bacillus subtilis levansucrase: a possible two step mechanism.. Eur J Biochem 163:379–387
    [Google Scholar]
  27. Petit-Glatron M. F., Grajcar L., Munz A., Chambert R. 1993; The contribution of the cell wall to a transmembrane calcium gradient could play a key role in Bacillus subtilis protein secretion.. Mol Microbiol 9:1097–1106
    [Google Scholar]
  28. Prats M., Tocanne J. F. 1986; Lateral proton conduction at lipid-water interfaces and its implications for the chemiosmotic-coupling hypothesis.. Nature 322:756–758
    [Google Scholar]
  29. Randall L.L., Hardy S.J.S, Tom J. R. 1987; Export of protein: a biochemical view.. Annu Rev Microbiol 41:507–541
    [Google Scholar]
  30. Sasamoto H., Nakasawa K., Tsutsumi K., Takase K., Yamane K. 1989; Signal peptide of Bacillus subtilis α-amylase.. J. Biochem 106:376–382
    [Google Scholar]
  31. Shioi J. I., Matsuura S., Imae Y. 1980; Quantitative measurements of proton motive force and motility in Bacillus subtilis. . J Bacteriol 144:891–897
    [Google Scholar]
  32. Simon S. M., Peskin C. S., Oster G. F. 1992; What drives the translocation of proteins?. Proc Natl Acad Sci USA 89:3770–3774
    [Google Scholar]
  33. Somogyi M. 1945; A new reagent for the determination of sugars.. J Biol Chem 160:61–73
    [Google Scholar]
  34. Stuart R. A., Cyr D. M., Craig E. A., Neupert W. 1994; Mitochondrial molecular chaperones; their role in protein translocation.. Trends Biochem Sci 19:87–92
    [Google Scholar]
  35. Teissi J., Gabriel B., Prats M. 1993; Lateral communication by fast proton conduction: a model membrane study.. Trends Biochem Sci 18243–246
    [Google Scholar]
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