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. 1996 Nov;8(11):2117–2126. doi: 10.1105/tpc.8.11.2117

Protein targeting and integration signal for the chloroplastic outer envelope membrane.

H M Li 1, L J Chen 1
PMCID: PMC161339  PMID: 8953775

Abstract

Most proteins in chloroplasts are encoded by the nuclear genome and synthesized in the cytosol. With the exception of most quter envelope membrane proteins, nuclear-encoded chloroplastic proteins are synthesized with N-terminal extensions that contain the chloroplast targeting information of these proteins. Most outer membrane proteins, however, are synthesized without extensions in the cytosol. Therefore, it is not clear where the chloroplastic outer membrane targeting information resides within these polypeptides. We have analyzed a chloroplastic outer membrane protein, OEP14 (outer envelope membrane protein of 14 kD, previously named OM14), and localized its outer membrane targeting and integration signal to the first 30 amino acids of the protein. This signal consists of a positively charged N-terminal portion followed by a hydrophobic core, bearing resemblance to the signal peptides of proteins targeted to the endoplasmic reticulum. However, a chimeric protein containing this signal fused to a passenger protein did not integrate into the endoplasmic reticulum membrane. Furthermore, membrane topology analysis indicated that the signal inserts into the chloroplastic outer membrane in an orientation opposite to that predicted by the "positive inside" rule.

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Selected References

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  1. Cline K., Werner-Washburne M., Andrews J., Keegstra K. Thermolysin is a suitable protease for probing the surface of intact pea chloroplasts. Plant Physiol. 1984 Jul;75(3):675–678. doi: 10.1104/pp.75.3.675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Eilers M., Hwang S., Schatz G. Unfolding and refolding of a purified precursor protein during import into isolated mitochondria. EMBO J. 1988 Apr;7(4):1139–1145. doi: 10.1002/j.1460-2075.1988.tb02923.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Endo T., Eilers M., Schatz G. Binding of a tightly folded artificial mitochondrial precursor protein to the mitochondrial outer membrane involves a lipid-mediated conformational change. J Biol Chem. 1989 Feb 15;264(5):2951–2956. [PubMed] [Google Scholar]
  4. Fischer K., Weber A., Arbinger B., Brink S., Eckerskorn C., Flügge U. I. The 24 kDa outer envelope membrane protein from spinach chloroplasts: molecular cloning, in vivo expression and import pathway of a protein with unusual properties. Plant Mol Biol. 1994 May;25(2):167–177. doi: 10.1007/BF00023235. [DOI] [PubMed] [Google Scholar]
  5. Gavel Y., Steppuhn J., Herrmann R., von Heijne G. The 'positive-inside rule' applies to thylakoid membrane proteins. FEBS Lett. 1991 Apr 22;282(1):41–46. doi: 10.1016/0014-5793(91)80440-e. [DOI] [PubMed] [Google Scholar]
  6. Hageman J., Baecke C., Ebskamp M., Pilon R., Smeekens S., Weisbeek P. Protein Import into and Sorting inside the Chloroplast Are Independent Processes. Plant Cell. 1990 May;2(5):479–494. doi: 10.1105/tpc.2.5.479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hartmann E., Rapoport T. A., Lodish H. F. Predicting the orientation of eukaryotic membrane-spanning proteins. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5786–5790. doi: 10.1073/pnas.86.15.5786. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hirsch S., Muckel E., Heemeyer F., von Heijne G., Soll J. A receptor component of the chloroplast protein translocation machinery. Science. 1994 Dec 23;266(5193):1989–1992. doi: 10.1126/science.7801125. [DOI] [PubMed] [Google Scholar]
  9. Keegstra K. Transport and routing of proteins into chloroplasts. Cell. 1989 Jan 27;56(2):247–253. doi: 10.1016/0092-8674(89)90898-2. [DOI] [PubMed] [Google Scholar]
  10. Keil P., Pfanner N. Insertion of MOM22 into the mitochondrial outer membrane strictly depends on surface receptors. FEBS Lett. 1993 Apr 26;321(2-3):197–200. doi: 10.1016/0014-5793(93)80107-6. [DOI] [PubMed] [Google Scholar]
  11. Kessler F., Blobel G., Patel H. A., Schnell D. J. Identification of two GTP-binding proteins in the chloroplast protein import machinery. Science. 1994 Nov 11;266(5187):1035–1039. doi: 10.1126/science.7973656. [DOI] [PubMed] [Google Scholar]
  12. Ko K., Bornemisza O., Kourtz L., Ko Z. W., Plaxton W. C., Cashmore A. R. Isolation and characterization of a cDNA clone encoding a cognate 70-kDa heat shock protein of the chloroplast envelope. J Biol Chem. 1992 Feb 15;267(5):2986–2993. [PubMed] [Google Scholar]
  13. Krishtalik L. I., Cramer W. A. On the physical basis for the cis-positive rule describing protein orientation in biological membranes. FEBS Lett. 1995 Aug 7;369(2-3):140–143. doi: 10.1016/0014-5793(95)00756-y. [DOI] [PubMed] [Google Scholar]
  14. Li H. M., Moore T., Keegstra K. Targeting of proteins to the outer envelope membrane uses a different pathway than transport into chloroplasts. Plant Cell. 1991 Jul;3(7):709–717. doi: 10.1105/tpc.3.7.709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Li H. M., Sullivan T. D., Keegstra K. Information for targeting to the chloroplastic inner envelope membrane is contained in the mature region of the maize Bt1-encoded protein. J Biol Chem. 1992 Sep 15;267(26):18999–19004. [PubMed] [Google Scholar]
  16. Lill R., Neupert W. Mechanisms of protein import across the mitochondrial outer membrane. Trends Cell Biol. 1996 Feb;6(2):56–61. doi: 10.1016/0962-8924(96)81015-4. [DOI] [PubMed] [Google Scholar]
  17. Markwell J., Bruce B. D., Keegstra K. Isolation of a carotenoid-containing sub-membrane particle from the chloroplastic envelope outer membrane of pea (Pisum sativum). J Biol Chem. 1992 Jul 15;267(20):13933–13937. [PubMed] [Google Scholar]
  18. Olsen L. J., Theg S. M., Selman B. R., Keegstra K. ATP is required for the binding of precursor proteins to chloroplasts. J Biol Chem. 1989 Apr 25;264(12):6724–6729. [PubMed] [Google Scholar]
  19. Perry S. E., Li H. M., Keegstra K. In vitro reconstitution of protein transport into chloroplasts. Methods Cell Biol. 1991;34:327–344. doi: 10.1016/s0091-679x(08)61688-x. [DOI] [PubMed] [Google Scholar]
  20. Salomon M., Fischer K., Flügge U. I., Soll J. Sequence analysis and protein import studies of an outer chloroplast envelope polypeptide. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5778–5782. doi: 10.1073/pnas.87.15.5778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Seedorf M., Waegemann K., Soll J. A constituent of the chloroplast import complex represents a new type of GTP-binding protein. Plant J. 1995 Mar;7(3):401–411. doi: 10.1046/j.1365-313x.1995.7030401.x. [DOI] [PubMed] [Google Scholar]
  23. Steger H. F., Söllner T., Kiebler M., Dietmeier K. A., Pfaller R., Trülzsch K. S., Tropschug M., Neupert W., Pfanner N. Import of ADP/ATP carrier into mitochondria: two receptors act in parallel. J Cell Biol. 1990 Dec;111(6 Pt 1):2353–2363. doi: 10.1083/jcb.111.6.2353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Söllner T., Griffiths G., Pfaller R., Pfanner N., Neupert W. MOM19, an import receptor for mitochondrial precursor proteins. Cell. 1989 Dec 22;59(6):1061–1070. doi: 10.1016/0092-8674(89)90762-9. [DOI] [PubMed] [Google Scholar]
  25. Söllner T., Pfaller R., Griffiths G., Pfanner N., Neupert W. A mitochondrial import receptor for the ADP/ATP carrier. Cell. 1990 Jul 13;62(1):107–115. doi: 10.1016/0092-8674(90)90244-9. [DOI] [PubMed] [Google Scholar]
  26. Tranel P. J., Froehlich J., Goyal A., Keegstra K. A component of the chloroplastic protein import apparatus is targeted to the outer envelope membrane via a novel pathway. EMBO J. 1995 Jun 1;14(11):2436–2446. doi: 10.1002/j.1460-2075.1995.tb07241.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Verner K., Schatz G. Import of an incompletely folded precursor protein into isolated mitochondria requires an energized inner membrane, but no added ATP. EMBO J. 1987 Aug;6(8):2449–2456. doi: 10.1002/j.1460-2075.1987.tb02524.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wu C., Ko K. Identification of an uncleavable targeting signal in the 70-kilodalton spinach chloroplast outer envelope membrane protein. J Biol Chem. 1993 Sep 15;268(26):19384–19391. [PubMed] [Google Scholar]
  29. van Loon A. P., Brändli A. W., Schatz G. The presequences of two imported mitochondrial proteins contain information for intracellular and intramitochondrial sorting. Cell. 1986 Mar 14;44(5):801–812. doi: 10.1016/0092-8674(86)90846-9. [DOI] [PubMed] [Google Scholar]
  30. von Heijne G., Gavel Y. Topogenic signals in integral membrane proteins. Eur J Biochem. 1988 Jul 1;174(4):671–678. doi: 10.1111/j.1432-1033.1988.tb14150.x. [DOI] [PubMed] [Google Scholar]
  31. von Heijne G., Steppuhn J., Herrmann R. G. Domain structure of mitochondrial and chloroplast targeting peptides. Eur J Biochem. 1989 Apr 1;180(3):535–545. doi: 10.1111/j.1432-1033.1989.tb14679.x. [DOI] [PubMed] [Google Scholar]
  32. von Heijne G. The signal peptide. J Membr Biol. 1990 May;115(3):195–201. doi: 10.1007/BF01868635. [DOI] [PubMed] [Google Scholar]

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