Abstract
Export through the cytoplasmic membrane and processing of the sak product in Escherichia coli cells were investigated with E. coli strains carrying pTS301, which produce large amounts of staphylokinase at 42 degrees C. High-level synthesis of the sak product caused transient accumulation not only of the staphylokinase precursor (pSAK) but also of the maltose-binding protein and outer membrane protein A precursors. Thus it was concluded that the sak product shares the export pathway with E. coli secreted proteins at least at a certain step. During high-level synthesis of the sak product, a significant amount of the newly synthesized pSAK remained unprocessed after a chase period, possibly causing the observed accumulation of pSAK. Accumulating pSAK did not mature for a long period, whereas the newly synthesized sak product was exclusively detected in the mature form. These results suggest that it is necessary for the sak product to enter the export pathway during or immediately after synthesis to be exported and processed normally.
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- 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]
- Blobel G., Dobberstein B. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma. J Cell Biol. 1975 Dec;67(3):835–851. doi: 10.1083/jcb.67.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Casadaban M. J. Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. J Mol Biol. 1976 Jul 5;104(3):541–555. doi: 10.1016/0022-2836(76)90119-4. [DOI] [PubMed] [Google Scholar]
- Chang C. N., Nielsen J. B., Izui K., Blobel G., Lampen J. O. Identification of the signal peptidase cleavage site in Bacillus licheniformis prepenicillinase. J Biol Chem. 1982 Apr 25;257(8):4340–4344. [PubMed] [Google Scholar]
- 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]
- Emr S. D., Bassford P. J., Jr Localization and processing of outer membrane and periplasmic proteins in Escherichia coli strains harboring export-specific suppressor mutations. J Biol Chem. 1982 May 25;257(10):5852–5860. [PubMed] [Google Scholar]
- Engelman D. M., Steitz T. A. The spontaneous insertion of proteins into and across membranes: the helical hairpin hypothesis. Cell. 1981 Feb;23(2):411–422. doi: 10.1016/0092-8674(81)90136-7. [DOI] [PubMed] [Google Scholar]
- Hayashi S., Chang S. Y., Chang S., Wu H. C. Modification and processing of Bacillus licheniformis prepenicillinase in Escherichia coli. Fate of mutant penicillinase lacking lipoprotein modification site. J Biol Chem. 1984 Aug 25;259(16):10448–10454. [PubMed] [Google Scholar]
- Inouye M., Halegoua S. Secretion and membrane localization of proteins in Escherichia coli. CRC Crit Rev Biochem. 1980;7(4):339–371. doi: 10.3109/10409238009105465. [DOI] [PubMed] [Google Scholar]
- 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]
- Josefsson L. G., Randall L. L. Different exported proteins in E. coli show differences in the temporal mode of processing in vivo. Cell. 1981 Jul;25(1):151–157. doi: 10.1016/0092-8674(81)90239-7. [DOI] [PubMed] [Google Scholar]
- 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]
- Kumamoto C. A., Beckwith J. Evidence for specificity at an early step in protein export in Escherichia coli. J Bacteriol. 1985 Jul;163(1):267–274. doi: 10.1128/jb.163.1.267-274.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Löfdahl S., Guss B., Uhlén M., Philipson L., Lindberg M. Gene for staphylococcal protein A. Proc Natl Acad Sci U S A. 1983 Feb;80(3):697–701. doi: 10.1073/pnas.80.3.697. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Minkley E. G., Jr Purification and characterization of pro-TraTp, the signal sequence-containing precursor of a secreted protein encoded by the F sex factor. J Bacteriol. 1984 May;158(2):464–473. doi: 10.1128/jb.158.2.464-473.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Morita T., Amemura M., Makino K., Shinagawa H., Magota K., Otsuji N., Nakata A. Hyperproduction of phosphate-binding protein, phoS, and pre-phoS proteins in Escherichia coli carrying a cloned phoS gene. Eur J Biochem. 1983 Feb 15;130(3):427–435. doi: 10.1111/j.1432-1033.1983.tb07169.x. [DOI] [PubMed] [Google Scholar]
- Oliver D. B., Beckwith J. E. coli mutant pleiotropically defective in the export of secreted proteins. Cell. 1981 Sep;25(3):765–772. doi: 10.1016/0092-8674(81)90184-7. [DOI] [PubMed] [Google Scholar]
- Osborn M. J., Gander J. E., Parisi E., Carson J. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Isolation and characterization of cytoplasmic and outer membrane. J Biol Chem. 1972 Jun 25;247(12):3962–3972. [PubMed] [Google Scholar]
- Pages J. M., Anba J., Bernadac A., Shinagawa H., Nakata A., Lazdunski C. Normal precursors of periplasmic proteins accumulated in the cytoplasm are not exported post-translationally in Escherichia coli. Eur J Biochem. 1984 Sep 17;143(3):499–505. doi: 10.1111/j.1432-1033.1984.tb08398.x. [DOI] [PubMed] [Google Scholar]
- Randall L. L. Translocation of domains of nascent periplasmic proteins across the cytoplasmic membrane is independent of elongation. Cell. 1983 May;33(1):231–240. doi: 10.1016/0092-8674(83)90352-5. [DOI] [PubMed] [Google Scholar]
- Russel M., Model P. Filamentous phage pre-coat is an integral membrane protein: analysis by a new method of membrane preparation. Cell. 1982 Jan;28(1):177–184. doi: 10.1016/0092-8674(82)90387-7. [DOI] [PubMed] [Google Scholar]
- Sako T. Overproduction of staphylokinase in Escherichia coli and its characterization. Eur J Biochem. 1985 Jun 18;149(3):557–563. doi: 10.1111/j.1432-1033.1985.tb08961.x. [DOI] [PubMed] [Google Scholar]
- Sako T., Sawaki S., Sakurai T., Ito S., Yoshizawa Y., Kondo I. Cloning and expression of the staphylokinase gene of Staphylococcus aureus in Escherichia coli. Mol Gen Genet. 1983;190(2):271–277. doi: 10.1007/BF00330650. [DOI] [PubMed] [Google Scholar]
- Sako T., Tsuchida N. Nucleotide sequence of the staphylokinase gene from Staphylococcus aureus. Nucleic Acids Res. 1983 Nov 25;11(22):7679–7693. doi: 10.1093/nar/11.22.7679. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sarvas M. O., Palva I. A. The penicillinase of Bacillus licheniformis is an outer membrane protein in Escherichia coli. J Bacteriol. 1983 Aug;155(2):657–663. doi: 10.1128/jb.155.2.657-663.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shiba K., Ito K., Yura T., Cerretti D. P. A defined mutation in the protein export gene within the spc ribosomal protein operon of Escherichia coli: isolation and characterization of a new temperature-sensitive secY mutant. EMBO J. 1984 Mar;3(3):631–635. doi: 10.1002/j.1460-2075.1984.tb01859.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Talmadge K., Stahl S., Gilbert W. Eukaryotic signal sequence transports insulin antigen in Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3369–3373. doi: 10.1073/pnas.77.6.3369. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tsukagoshi N., Ihara H., Yamagata H., Udaka S. Cloning and expression of a thermophilic alpha-amylase gene from Bacillus stearothermophilus in Escherichia coli. Mol Gen Genet. 1984;193(1):58–63. doi: 10.1007/BF00327414. [DOI] [PubMed] [Google Scholar]
- Wickner W. The assembly of proteins into biological membranes: The membrane trigger hypothesis. Annu Rev Biochem. 1979;48:23–45. doi: 10.1146/annurev.bi.48.070179.000323. [DOI] [PubMed] [Google Scholar]
- Zemel-Dreasen O., Zamir A. Secretion and processing of an immunoglobulin light chain in Escherichia coli. Gene. 1984 Mar;27(3):315–322. doi: 10.1016/0378-1119(84)90076-3. [DOI] [PubMed] [Google Scholar]
- von Heijne G., Blomberg C. Trans-membrane translocation of proteins. The direct transfer model. Eur J Biochem. 1979 Jun;97(1):175–181. doi: 10.1111/j.1432-1033.1979.tb13100.x. [DOI] [PubMed] [Google Scholar]