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. 1986 Jun;166(3):728–732. doi: 10.1128/jb.166.3.728-732.1986

Excretion of the penicillinase of an alkalophilic Bacillus sp. through the Escherichia coli outer membrane is caused by insertional activation of the kil gene in plasmid pMB9.

T Kobayashi, C Kato, T Kudo, K Horikoshi
PMCID: PMC215185  PMID: 3011739

Abstract

Most of the cloned penicillinase from alkalophilic Bacillus sp. strain 170 and alkaline phosphatase were released into the culture medium by Escherichia coli strains bearing plasmid pEAP1 or pEAP2 (T. Kudo, C. Kato, and K. Horikoshi, J. Bacteriol. 156:949-951, 1983). We analyzed the basis for excretion of periplasmic enzymes in the cells bearing these plasmids. Several experiments such as subcloning, insertion of a chloramphenicol acetyltransferase cartridge, and DNA sequencing were done. A dormant kil gene in plasmid pMB9 was expressed by a promoter of the inserted DNA fragment of alkalophilic Bacillus sp. strain 170, and as a result, the outer membrane of E. coli became permeable, allowing the proteins to be excreted without cell lysis.

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Selected References

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  1. Bolivar F., Rodriguez R. L., Betlach M. C., Boyer H. W. Construction and characterization of new cloning vehicles. I. Ampicillin-resistant derivatives of the plasmid pMB9. Gene. 1977;2(2):75–93. doi: 10.1016/0378-1119(77)90074-9. [DOI] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  3. Close T. J., Rodriguez R. L. Construction and characterization of the chloramphenicol-resistance gene cartridge: a new approach to the transcriptional mapping of extrachromosomal elements. Gene. 1982 Dec;20(2):305–316. doi: 10.1016/0378-1119(82)90048-8. [DOI] [PubMed] [Google Scholar]
  4. Ebina Y., Nakazawa A. Cyclic AMP-dependent initiation and rho-dependent termination of colicin E1 gene transcription. J Biol Chem. 1983 Jun 10;258(11):7072–7078. [PubMed] [Google Scholar]
  5. Gilman M. Z., Chamberlin M. J. Developmental and genetic regulation of Bacillus subtilis genes transcribed by sigma 28-RNA polymerase. Cell. 1983 Nov;35(1):285–293. doi: 10.1016/0092-8674(83)90231-3. [DOI] [PubMed] [Google Scholar]
  6. Goldfarb D. S., Rodriguez R. L., Doi R. H. Translational block to expression of the Escherichia coli Tn9-derived chloramphenicol-resistance gene in Bacillus subtilis. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5886–5890. doi: 10.1073/pnas.79.19.5886. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hakkaart M. J., Veltkamp E., Nijkamp H. J. Protein H encoded by plasmid Clo DF13 involved in lysis of the bacterial host. I. Localisation of the gene and identification and subcellular localisation of the gene H product. Mol Gen Genet. 1981;183(2):318–325. doi: 10.1007/BF00270635. [DOI] [PubMed] [Google Scholar]
  8. Kudo T., Kato C., Horikoshi K. Excretion of the penicillinase of an alkalophilic Bacillus sp. through the Escherichia coli outer membrane. J Bacteriol. 1983 Nov;156(2):949–951. doi: 10.1128/jb.156.2.949-951.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kudo T., Yoshitake J., Kato C., Usami R., Horikoshi K. Cloning of a developmentally regulated element from alkalophilic Bacillus subtilis DNA. J Bacteriol. 1985 Jan;161(1):158–163. doi: 10.1128/jb.161.1.158-163.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  11. Nakamura K., Inouye M. DNA sequence of the gene for the outer membrane lipoprotein of E. coli: an extremely AT-rich promoter. Cell. 1979 Dec;18(4):1109–1117. doi: 10.1016/0092-8674(79)90224-1. [DOI] [PubMed] [Google Scholar]
  12. Oka A., Nomura N., Morita M., Sugisaki H., Sugimoto K., Takanami M. Nucleotide sequence of small ColE1 derivatives: structure of the regions essential for autonomous replication and colicin E1 immunity. Mol Gen Genet. 1979 May 4;172(2):151–159. doi: 10.1007/BF00268276. [DOI] [PubMed] [Google Scholar]
  13. Oudega B., Stegehuis F., van Tiel-Menkveld G. J., de Graaf F. K. Protein H encoded by plasmid CloDF13 is involved in excretion of cloacin DF13. J Bacteriol. 1982 Jun;150(3):1115–1121. doi: 10.1128/jb.150.3.1115-1121.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Pugsley A. P., Schwartz M. A genetic approach to the study of mitomycin-induced lysis of Escherichia coli K-12 strains which produce colicin E2. Mol Gen Genet. 1983;190(3):366–372. doi: 10.1007/BF00331060. [DOI] [PubMed] [Google Scholar]
  15. Pugsley A. P., Schwartz M. Colicin E2 release: lysis, leakage or secretion? Possible role of a phospholipase. EMBO J. 1984 Oct;3(10):2393–2397. doi: 10.1002/j.1460-2075.1984.tb02145.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Pugsley A. P. The ins and outs of colicins. Part I: Production, and translocation across membranes. Microbiol Sci. 1984 Oct;1(7):168–175. [PubMed] [Google Scholar]
  17. Sabik J. F., Suit J. L., Luria S. E. cea-kil operon of the ColE1 plasmid. J Bacteriol. 1983 Mar;153(3):1479–1485. doi: 10.1128/jb.153.3.1479-1485.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sargent M. G. Rapid fixed-time assay for penicillinase. J Bacteriol. 1968 Apr;95(4):1493–1494. doi: 10.1128/jb.95.4.1493-1494.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sawai T., Takahashi I., Yamagishi S. Iodometric assay method for beta-lactamase with various beta-lactam antibiotics as substrates. Antimicrob Agents Chemother. 1978 Jun;13(6):910–913. doi: 10.1128/aac.13.6.910. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Shaw W. V. Chloramphenicol acetyltransferase from chloramphenicol-resistant bacteria. Methods Enzymol. 1975;43:737–755. doi: 10.1016/0076-6879(75)43141-x. [DOI] [PubMed] [Google Scholar]
  22. Yamada M., Nakazawa A. Factors necessary for the export process of colicin E1 across cytoplasmic membrane of Escherichia coli. Eur J Biochem. 1984 Apr 16;140(2):249–255. doi: 10.1111/j.1432-1033.1984.tb08095.x. [DOI] [PubMed] [Google Scholar]

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