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. 1988 Oct;170(10):4445–4450. doi: 10.1128/jb.170.10.4445-4450.1988

Two regions of mature periplasmic maltose-binding protein of Escherichia coli involved in secretion.

P Duplay 1, M Hofnung 1
PMCID: PMC211475  PMID: 3049532

Abstract

Six mutations in malE, the structural gene for the periplasmic maltose-binding protein (MBP) from Escherichia coli, prevent growth on maltose as a carbon source, as well as release of the mutant proteins by the cold osmotic-shock procedure. These mutations correspond to insertion of an oligonucleotide linker, concomitant with a deletion. One of the mutations (malE127) affects the N-terminal extension (the signal peptide), whereas the five others lie within the mature protein. As expected, the export of protein MalE127 is blocked at an early stage. This protein is neither processed to maturity nor sensitive to proteinase K in spheroplasts. In contrast, in the five other mutants, the signal peptide is cleaved and the protein is accessible to proteinase K added to spheroplasts. This indicates that the five mutant proteins are, at least in part, exported through the inner membrane. We propose that the corresponding mutations define two regions of the mature protein (between residues 18 and 42 and between residues 280 and 306), which are important for release of the protein from the inner membrane into the periplasm. We discuss the results in terms of possible conformational changes at this late step of export to the periplasm.

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Selected References

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

  1. Bankaitis V. A., Bassford P. J., Jr Sequences within the mature maltose-binding protein of Escherichia coli may be actively involved in initiating the export process. Ann Inst Pasteur Microbiol. 1985 Jul-Aug;136B(1):3–7. doi: 10.1016/s0769-2609(85)80001-6. [DOI] [PubMed] [Google Scholar]
  2. Bankaitis V. A., Bassford P. J., Jr The synthesis of export-defective proteins can interfere with normal protein export in Escherichia coli. J Biol Chem. 1984 Oct 10;259(19):12193–12200. [PubMed] [Google Scholar]
  3. Bankaitis V. A., Rasmussen B. A., Bassford P. J., Jr Intragenic suppressor mutations that restore export of maltose binding protein with a truncated signal peptide. Cell. 1984 May;37(1):243–252. doi: 10.1016/0092-8674(84)90320-9. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Bedouelle H., Bassford P. J., Jr, Fowler A. V., Zabin I., Beckwith J., Hofnung M. Mutations which alter the function of the signal sequence of the maltose binding protein of Escherichia coli. Nature. 1980 May 8;285(5760):78–81. doi: 10.1038/285078a0. [DOI] [PubMed] [Google Scholar]
  6. Bedouelle H., Duplay P. Production in Escherichia coli and one-step purification of bifunctional hybrid proteins which bind maltose. Export of the Klenow polymerase into the periplasmic space. Eur J Biochem. 1988 Feb 1;171(3):541–549. doi: 10.1111/j.1432-1033.1988.tb13823.x. [DOI] [PubMed] [Google Scholar]
  7. Brass J. M., Manson M. D. Reconstitution of maltose chemotaxis in Escherichia coli by addition of maltose-binding protein to calcium-treated cells of maltose regulon mutants. J Bacteriol. 1984 Mar;157(3):881–890. doi: 10.1128/jb.157.3.881-890.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cover W. H., Ryan J. P., Bassford P. J., Jr, Walsh K. A., Bollinger J., Randall L. L. Suppression of a signal sequence mutation by an amino acid substitution in the mature portion of the maltose-binding protein. J Bacteriol. 1987 May;169(5):1794–1800. doi: 10.1128/jb.169.5.1794-1800.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dalbey R. E., Wickner W. Leader peptidase catalyzes the release of exported proteins from the outer surface of the Escherichia coli plasma membrane. J Biol Chem. 1985 Dec 15;260(29):15925–15931. [PubMed] [Google Scholar]
  10. Davis N. G., Model P. An artificial anchor domain: hydrophobicity suffices to stop transfer. Cell. 1985 Jun;41(2):607–614. doi: 10.1016/s0092-8674(85)80033-7. [DOI] [PubMed] [Google Scholar]
  11. Dierstein R., Wickner W. The leader region of pre-maltose binding protein binds amphiphiles. A model for self-assembly in protein export. J Biol Chem. 1985 Dec 15;260(29):15919–15924. [PubMed] [Google Scholar]
  12. Duplay P., Szmelcman S., Bedouelle H., Hofnung M. Silent and functional changes in the periplasmic maltose-binding protein of Escherichia coli K12. I. Transport of maltose. J Mol Biol. 1987 Apr 20;194(4):663–673. doi: 10.1016/0022-2836(87)90243-9. [DOI] [PubMed] [Google Scholar]
  13. Ferenci T., Randall L. L. Precursor maltose-binding protein is active in binding substrate. J Biol Chem. 1979 Oct 25;254(20):9979–9981. [PubMed] [Google Scholar]
  14. Fikes J. D., Bankaitis V. A., Ryan J. P., Bassford P. J., Jr Mutational alterations affecting the export competence of a truncated but fully functional maltose-binding protein signal peptide. J Bacteriol. 1987 Jun;169(6):2345–2351. doi: 10.1128/jb.169.6.2345-2351.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fikes J. D., Bassford P. J., Jr Export of unprocessed precursor maltose-binding protein to the periplasm of Escherichia coli cells. J Bacteriol. 1987 Jun;169(6):2352–2359. doi: 10.1128/jb.169.6.2352-2359.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fitts R., Reuveny Z., van Amsterdam J., Mulholland J., Botstein D. Substitution of tyrosine for either cysteine in beta-lactamase prevents release from the membrane during secretion. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8540–8543. doi: 10.1073/pnas.84.23.8540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hengge R., Boos W. Defective secretion of maltose- and ribose-binding proteins caused by a truncated periplasmic protein in Escherichia coli. J Bacteriol. 1985 Jun;162(3):972–978. doi: 10.1128/jb.162.3.972-978.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ito K., Bassford P. J., Jr, Beckwith J. Protein localization in E. coli: is there a common step in the secretion of periplasmic and outer-membrane proteins? Cell. 1981 Jun;24(3):707–717. doi: 10.1016/0092-8674(81)90097-0. [DOI] [PubMed] [Google Scholar]
  19. Ito K., Beckwith J. R. Role of the mature protein sequence of maltose-binding protein in its secretion across the E. coli cytoplasmic membrane. Cell. 1981 Jul;25(1):143–150. doi: 10.1016/0092-8674(81)90238-5. [DOI] [PubMed] [Google Scholar]
  20. Ito K. Purification of the precursor form of maltose-binding protein, a periplasmic protein of Escherichia coli. J Biol Chem. 1982 Sep 10;257(17):9895–9897. [PubMed] [Google Scholar]
  21. Jacobson G. R., Rosenbusch J. P. Abundance and membrane association of elongation factor Tu in E. coli. Nature. 1976 May 6;261(5555):23–26. doi: 10.1038/261023a0. [DOI] [PubMed] [Google Scholar]
  22. Jacobson G. R., Takacs B. J., Rosenbusch J. P. Properties of a major protein released from Escherichia coli by osmotic shock. Biochemistry. 1976 Jun 1;15(11):2297–2303. doi: 10.1021/bi00656a008. [DOI] [PubMed] [Google Scholar]
  23. Koshland D., Botstein D. Evidence for posttranslational translocation of beta-lactamase across the bacterial inner membrane. Cell. 1982 Oct;30(3):893–902. doi: 10.1016/0092-8674(82)90294-x. [DOI] [PubMed] [Google Scholar]
  24. Kuhn A., Wickner W. Isolation of mutants in M13 coat protein that affect its synthesis, processing, and assembly into phage. J Biol Chem. 1985 Dec 15;260(29):15907–15913. [PubMed] [Google Scholar]
  25. 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]
  26. Lin J. J., Kanazawa H., Wu H. C. Assembly of outer membrane lipoprotein in an Escherichia coli mutant with a single amino acid replacement within the signal sequence of prolipoprotein. J Bacteriol. 1980 Feb;141(2):550–557. doi: 10.1128/jb.141.2.550-557.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Minsky A., Summers R. G., Knowles J. R. Secretion of beta-lactamase into the periplasm of Escherichia coli: evidence for a distinct release step associated with a conformational change. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4180–4184. doi: 10.1073/pnas.83.12.4180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Randall L. L. Function of protonmotive force in translocation of protein across membranes. Methods Enzymol. 1986;125:129–138. doi: 10.1016/s0076-6879(86)25012-0. [DOI] [PubMed] [Google Scholar]
  29. Randall L. L., Hardy S. J. Correlation of competence for export with lack of tertiary structure of the mature species: a study in vivo of maltose-binding protein in E. coli. Cell. 1986 Sep 12;46(6):921–928. doi: 10.1016/0092-8674(86)90074-7. [DOI] [PubMed] [Google Scholar]
  30. Randall L. L., Hardy S. J., Thom J. R. Export of protein: a biochemical view. Annu Rev Microbiol. 1987;41:507–541. doi: 10.1146/annurev.mi.41.100187.002451. [DOI] [PubMed] [Google Scholar]
  31. Ray P., Dev I., MacGregor C., Bassford P., Jr Signal peptidases. Curr Top Microbiol Immunol. 1986;125:75–102. doi: 10.1007/978-3-642-71251-7_7. [DOI] [PubMed] [Google Scholar]
  32. Ryan J. P., Duncan M. C., Bankaitis V. A., Bassford P. J., Jr Intragenic reversion mutations that improve export of maltose-binding protein in Escherichia coli malE signal sequence mutants. J Biol Chem. 1986 Mar 5;261(7):3389–3395. [PubMed] [Google Scholar]
  33. Wolfe P. B., Wickner W., Goodman J. M. Sequence of the leader peptidase gene of Escherichia coli and the orientation of leader peptidase in the bacterial envelope. J Biol Chem. 1983 Oct 10;258(19):12073–12080. [PubMed] [Google Scholar]
  34. von Heijne G. Signal sequences. The limits of variation. J Mol Biol. 1985 Jul 5;184(1):99–105. doi: 10.1016/0022-2836(85)90046-4. [DOI] [PubMed] [Google Scholar]

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