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. 1991 Jan;173(1):276–282. doi: 10.1128/jb.173.1.276-282.1991

Effect of signal sequence alterations on export of levansucrase in Bacillus subtilis.

T V Borchert 1, V Nagarajan 1
PMCID: PMC207184  PMID: 1898923

Abstract

A series of alterations in the Bacillus amyloliquefaciens levansucrase signal peptide were made by in vitro mutagenesis, and their effect on the secretion of levansucrase in Bacillus subtilis was studied. Some of the alterations resulted in a completely defective signal peptide. These included the removal of positively charged residues from the N-terminus and disruption of the hydrophobic core of the signal peptide either by introducing a charged residue or by deleting five or more amino acids. Analysis of the signal peptide processing-site alterations revealed that small residues are preferred at the -1 and -3 positions. However, a wide variety of amino acids are tolerated at the +1 position.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Beckwith J., Ferro-Novick S. Genetic studies on protein export in bacteria. Curr Top Microbiol Immunol. 1986;125:5–27. doi: 10.1007/978-3-642-71251-7_2. [DOI] [PubMed] [Google Scholar]
  2. Briggs M. S., Gierasch L. M. Molecular mechanisms of protein secretion: the role of the signal sequence. Adv Protein Chem. 1986;38:109–180. doi: 10.1016/s0065-3233(08)60527-6. [DOI] [PubMed] [Google Scholar]
  3. Chou P. Y., Fasman G. D. Empirical predictions of protein conformation. Annu Rev Biochem. 1978;47:251–276. doi: 10.1146/annurev.bi.47.070178.001343. [DOI] [PubMed] [Google Scholar]
  4. Emr S. D., Hanley-Way S., Silhavy T. J. Suppressor mutations that restore export of a protein with a defective signal sequence. Cell. 1981 Jan;23(1):79–88. doi: 10.1016/0092-8674(81)90272-5. [DOI] [PubMed] [Google Scholar]
  5. Emr S. D., Silhavy T. J. Importance of secondary structure in the signal sequence for protein secretion. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4599–4603. doi: 10.1073/pnas.80.15.4599. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fikes J. D., Barkocy-Gallagher G. A., Klapper D. G., Bassford P. J., Jr Maturation of Escherichia coli maltose-binding protein by signal peptidase I in vivo. Sequence requirements for efficient processing and demonstration of an alternate cleavage site. J Biol Chem. 1990 Feb 25;265(6):3417–3423. [PubMed] [Google Scholar]
  7. Gierasch L. M. Signal sequences. Biochemistry. 1989 Feb 7;28(3):923–930. doi: 10.1021/bi00429a001. [DOI] [PubMed] [Google Scholar]
  8. Iino T., Takahashi M., Sako T. Role of amino-terminal positive charge on signal peptide in staphylokinase export across the cytoplasmic membrane of Escherichia coli. J Biol Chem. 1987 May 25;262(15):7412–7417. [PubMed] [Google Scholar]
  9. Inouye S., Soberon X., Franceschini T., Nakamura K., Itakura K., Inouye M. Role of positive charge on the amino-terminal region of the signal peptide in protein secretion across the membrane. Proc Natl Acad Sci U S A. 1982 Jun;79(11):3438–3441. doi: 10.1073/pnas.79.11.3438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kadonaga J. T., Gautier A. E., Straus D. R., Charles A. D., Edge M. D., Knowles J. R. The role of the beta-lactamase signal sequence in the secretion of proteins by Escherichia coli. J Biol Chem. 1984 Feb 25;259(4):2149–2154. [PubMed] [Google Scholar]
  11. Kuhn A., Wickner W. Conserved residues of the leader peptide are essential for cleavage by leader peptidase. J Biol Chem. 1985 Dec 15;260(29):15914–15918. [PubMed] [Google Scholar]
  12. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  13. Lehnhardt S., Pollitt S., Inouye M. The differential effect on two hybrid proteins of deletion mutations within the hydrophobic region of the Escherichia coli OmpA signal peptide. J Biol Chem. 1987 Feb 5;262(4):1716–1719. [PubMed] [Google Scholar]
  14. Michaelis S., Hunt J. F., Beckwith J. Effects of signal sequence mutations on the kinetics of alkaline phosphatase export to the periplasm in Escherichia coli. J Bacteriol. 1986 Jul;167(1):160–167. doi: 10.1128/jb.167.1.160-167.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Murén E. M., Randall L. L. Export of alpha-amylase by Bacillus amyloliquefaciens requires proton motive force. J Bacteriol. 1985 Nov;164(2):712–716. doi: 10.1128/jb.164.2.712-716.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Nagarajan V. System for secretion of heterologous proteins in Bacillus subtilis. Methods Enzymol. 1990;185:214–223. doi: 10.1016/0076-6879(90)85021-f. [DOI] [PubMed] [Google Scholar]
  17. Nakamura K., Fujita Y., Itoh Y., Yamane K. Modification of length, hydrophobic properties and electric charge of Bacillus subtilis alpha-amylase signal peptide and their different effects on the production of secretory proteins in B. subtilis and Escherichia coli cells. Mol Gen Genet. 1989 Mar;216(1):1–9. doi: 10.1007/BF00332223. [DOI] [PubMed] [Google Scholar]
  18. Paddon C. J., Vasantha N., Hartley R. W. Translation and processing of Bacillus amyloliquefaciens extracellular RNase. J Bacteriol. 1989 Feb;171(2):1185–1187. doi: 10.1128/jb.171.2.1185-1187.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Palva I., Sarvas M., Lehtovaara P., Sibakov M., Käriäinen L. Secretion of Escherichia coli beta-lactamase from Bacillus subtilis by the aid of alpha-amylase signal sequence. Proc Natl Acad Sci U S A. 1982 Sep;79(18):5582–5586. doi: 10.1073/pnas.79.18.5582. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Perlman D., Halvorson H. O. A putative signal peptidase recognition site and sequence in eukaryotic and prokaryotic signal peptides. J Mol Biol. 1983 Jun 25;167(2):391–409. doi: 10.1016/s0022-2836(83)80341-6. [DOI] [PubMed] [Google Scholar]
  21. Petit-Glatron M. F., Benyahia F., Chambert R. Secretion of Bacillus subtilis levansucrase: a possible two-step mechanism. Eur J Biochem. 1987 Mar 2;163(2):379–387. doi: 10.1111/j.1432-1033.1987.tb10810.x. [DOI] [PubMed] [Google Scholar]
  22. Puziss J. W., Fikes J. D., Bassford P. J., Jr Analysis of mutational alterations in the hydrophilic segment of the maltose-binding protein signal peptide. J Bacteriol. 1989 May;171(5):2303–2311. doi: 10.1128/jb.171.5.2303-2311.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stader J., Gansheroff L. J., Silhavy T. J. New suppressors of signal-sequence mutations, prlG, are linked tightly to the secE gene of Escherichia coli. Genes Dev. 1989 Jul;3(7):1045–1052. doi: 10.1101/gad.3.7.1045. [DOI] [PubMed] [Google Scholar]
  24. Takase K., Mizuno H., Yamane K. NH2-terminal processing of Bacillus subtilis alpha-amylase. J Biol Chem. 1988 Aug 15;263(23):11548–11553. [PubMed] [Google Scholar]
  25. Vasantha N., Thompson L. D., Rhodes C., Banner C., Nagle J., Filpula D. Genes for alkaline protease and neutral protease from Bacillus amyloliquefaciens contain a large open reading frame between the regions coding for signal sequence and mature protein. J Bacteriol. 1984 Sep;159(3):811–819. doi: 10.1128/jb.159.3.811-819.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. von Heijne G. A new method for predicting signal sequence cleavage sites. Nucleic Acids Res. 1986 Jun 11;14(11):4683–4690. doi: 10.1093/nar/14.11.4683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. von Heijne G. Patterns of amino acids near signal-sequence cleavage sites. Eur J Biochem. 1983 Jun 1;133(1):17–21. doi: 10.1111/j.1432-1033.1983.tb07424.x. [DOI] [PubMed] [Google Scholar]

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