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. 1992 Apr;174(7):2095–2101. doi: 10.1128/jb.174.7.2095-2101.1992

Structural determinants in addition to the amino-terminal sorting sequence influence membrane localization of Escherichia coli lipoproteins.

J M Gennity 1, H Kim 1, M Inouye 1
PMCID: PMC205825  PMID: 1551832

Abstract

The lipid-modified nine-residue amino-terminal sequence of the mature form of the major outer membrane lipoprotein of Escherichia coli contains information that is responsible for sorting to either the inner or outer membrane. Fusion of this sorting sequence to beta-lactamase is sufficient for localization of the resultant lipo-beta-lactamase to the outer membrane (J. Ghrayeb and M. Inouye, J. Biol. Chem. 259:463-467, 1984). Substitution of the serine adjacent to the amino-terminal lipid-modified cysteine residue of the sorting sequence with the negatively charged residue aspartate causes inner membrane localization (K. Yamaguchi, F. Yu, and M. Inouye, Cell 53:423-432, 1988). Fusion of the aspartate-containing nine-residue inner membrane localization signal to the normally outer membrane lipoprotein bacteriocin release protein does cause partial localization to the inner membrane. However, a single replacement of the glutamine adjacent to the amino-terminal lipid-modified cysteine residue of bacteriocin release protein with aspartate causes no inner membrane localization. Therefore, an aspartate residue itself lacks the information necessary for inner membrane sorting when removed from the structural context provided by the additional eight residues of the sorting sequence. Although the aspartate-containing inner membrane sorting sequence causes an almost quantitative localization to the inner membrane when fused to the otherwise soluble protein beta-lactamase, this sequence cannot prevent significant outer membrane localization when fused to proteins (bacteriocin release protein and OmpA) normally found in the outer membrane. Therefore, structural determinants in addition to the amino-terminal sorting sequence influence the membrane localization of lipoproteins.

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Selected References

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

  1. De Graaf F. K., Oudega B. Production and release of cloacin DF13 and related colicins. Curr Top Microbiol Immunol. 1986;125:183–205. doi: 10.1007/978-3-642-71251-7_11. [DOI] [PubMed] [Google Scholar]
  2. Dornmair K., Kiefer H., Jähnig F. Refolding of an integral membrane protein. OmpA of Escherichia coli. J Biol Chem. 1990 Nov 5;265(31):18907–18911. [PubMed] [Google Scholar]
  3. Emory S. A., Belasco J. G. The ompA 5' untranslated RNA segment functions in Escherichia coli as a growth-rate-regulated mRNA stabilizer whose activity is unrelated to translational efficiency. J Bacteriol. 1990 Aug;172(8):4472–4481. doi: 10.1128/jb.172.8.4472-4481.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Filip C., Fletcher G., Wulff J. L., Earhart C. F. Solubilization of the cytoplasmic membrane of Escherichia coli by the ionic detergent sodium-lauryl sarcosinate. J Bacteriol. 1973 Sep;115(3):717–722. doi: 10.1128/jb.115.3.717-722.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Freudl R., Klose M., Henning U. Export and sorting of the Escherichia coli outer membrane protein OmpA. J Bioenerg Biomembr. 1990 Jun;22(3):441–449. doi: 10.1007/BF00763176. [DOI] [PubMed] [Google Scholar]
  6. Freudl R., Schwarz H., Stierhof Y. D., Gamon K., Hindennach I., Henning U. An outer membrane protein (OmpA) of Escherichia coli K-12 undergoes a conformational change during export. J Biol Chem. 1986 Aug 25;261(24):11355–11361. [PubMed] [Google Scholar]
  7. Gennity J. M., Inouye M. The protein sequence responsible for lipoprotein membrane localization in Escherichia coli exhibits remarkable specificity. J Biol Chem. 1991 Sep 5;266(25):16458–16464. [PubMed] [Google Scholar]
  8. Ghrayeb J., Inouye M. Nine amino acid residues at the NH2-terminal of lipoprotein are sufficient for its modification, processing, and localization in the outer membrane of Escherichia coli. J Biol Chem. 1984 Jan 10;259(1):463–467. [PubMed] [Google Scholar]
  9. Klose M., Schwarz H., MacIntyre S., Freudl R., Eschbach M. L., Henning U. Internal deletions in the gene for an Escherichia coli outer membrane protein define an area possibly important for recognition of the outer membrane by this polypeptide. J Biol Chem. 1988 Sep 15;263(26):13291–13296. [PubMed] [Google Scholar]
  10. Klose M., Schwarz H., MacIntyre S., Freudl R., Eschbach M. L., Henning U. Internal deletions in the gene for an Escherichia coli outer membrane protein define an area possibly important for recognition of the outer membrane by this polypeptide. J Biol Chem. 1988 Sep 15;263(26):13291–13296. [PubMed] [Google Scholar]
  11. 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]
  12. Lopilato J., Bortner S., Beckwith J. Mutations in a new chromosomal gene of Escherichia coli K-12, pcnB, reduce plasmid copy number of pBR322 and its derivatives. Mol Gen Genet. 1986 Nov;205(2):285–290. doi: 10.1007/BF00430440. [DOI] [PubMed] [Google Scholar]
  13. Luirink J., Duim B., de Gier J. W., Oudega B. Functioning of the stable signal peptide of the pCloDF13-encoded bacteriocin release protein. Mol Microbiol. 1991 Feb;5(2):393–399. doi: 10.1111/j.1365-2958.1991.tb02121.x. [DOI] [PubMed] [Google Scholar]
  14. Luirink J., Watanabe T., Wu H. C., Stegehuis F., de Graaf F. K., Oudega B. Modification, processing, and subcellular localization in Escherichia coli of the pCloDF13-encoded bacteriocin release protein fused to the mature portion of beta-lactamase. J Bacteriol. 1987 May;169(5):2245–2250. doi: 10.1128/jb.169.5.2245-2250.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Nakamura K., Inouye M. Construction of versatile expression cloning vehicles using the lipoprotein gene of Escherichia coli. EMBO J. 1982;1(6):771–775. doi: 10.1002/j.1460-2075.1982.tb01244.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Pugsley A. P., Kornacker M. G. Secretion of the cell surface lipoprotein pullulanase in Escherichia coli. Cooperation or competition between the specific secretion pathway and the lipoprotein sorting pathway. J Biol Chem. 1991 Jul 25;266(21):13640–13645. [PubMed] [Google Scholar]
  17. Schweizer M., Hindennach I., Garten W., Henning U. Major proteins of the Escherichia coli outer cell envelope membrane. Interaction of protein II with lipopolysaccharide. Eur J Biochem. 1978 Jan 2;82(1):211–217. doi: 10.1111/j.1432-1033.1978.tb12013.x. [DOI] [PubMed] [Google Scholar]
  18. Schägger H., von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal Biochem. 1987 Nov 1;166(2):368–379. doi: 10.1016/0003-2697(87)90587-2. [DOI] [PubMed] [Google Scholar]
  19. Tanji Y., Gennity J., Pollitt S., Inouye M. Effect of OmpA signal peptide mutations on OmpA secretion, synthesis, and assembly. J Bacteriol. 1991 Mar;173(6):1997–2005. doi: 10.1128/jb.173.6.1997-2005.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Vogel H., Jähnig F. Models for the structure of outer-membrane proteins of Escherichia coli derived from raman spectroscopy and prediction methods. J Mol Biol. 1986 Jul 20;190(2):191–199. doi: 10.1016/0022-2836(86)90292-5. [DOI] [PubMed] [Google Scholar]
  21. Yamaguchi K., Yu F., Inouye M. A single amino acid determinant of the membrane localization of lipoproteins in E. coli. Cell. 1988 May 6;53(3):423–432. doi: 10.1016/0092-8674(88)90162-6. [DOI] [PubMed] [Google Scholar]

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