Skip to main content
NCBI home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1995 Apr;61(4):1610–1613. doi: 10.1128/aem.61.4.1610-1613.1995

Expression and secretion of heterologous proteases by Corynebacterium glutamicum.

H Billman-Jacobe 1, L Wang 1, A Kortt 1, D Stewart 1, A Radford 1
PMCID: PMC167416  PMID: 7747974

Abstract

Genes encoding the basic protease of Dichelobacter nodosus (bprV) and the subtilisin of Bacillus subtilis (aprE) were cloned and expressed in Corynebacterium glutamicum. In each case, enzymatically active protein was detected in the supernatants of liquid cultures. While the secretion of subtilisin was directed by its own signal peptide, the natural signal peptide of the bprV basic protease did not facilitate secretion. A hybrid aprE-bprV gene in which the promoter and signal peptide coding sequences of subtilisin replaced those of bprV could be expressed, and basic protease was secreted by C. glutamicum. Expression of these proteases in C. glutamicum provides an opportunity to compare protein secretion from this gram-positive host with that from other gram-positive and gram-negative bacteria.

Full Text

The Full Text of this article is available as a PDF (251.7 KB).

Selected References

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

  1. Billman-Jacobe H., Hodgson A. L., Lightowlers M., Wood P. R., Radford A. J. Expression of ovine gamma interferon in Escherichia coli and Corynebacterium glutamicum. Appl Environ Microbiol. 1994 May;60(5):1641–1645. doi: 10.1128/aem.60.5.1641-1645.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chang S. Engineering for protein secretion in gram-positive bacteria. Methods Enzymol. 1987;153:507–516. doi: 10.1016/0076-6879(87)53075-0. [DOI] [PubMed] [Google Scholar]
  3. Kortt A. A., O'Donnell I. J., Stewart D. J., Clark B. L. Activities and partial purification of extracellular proteases of Bacteroides nodosus from virulent and benign footrot. Aust J Biol Sci. 1982;35(5):481–489. doi: 10.1071/bi9820481. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Liebl W., Sinskey A. J., Schleifer K. H. Expression, secretion, and processing of staphylococcal nuclease by Corynebacterium glutamicum. J Bacteriol. 1992 Mar;174(6):1854–1861. doi: 10.1128/jb.174.6.1854-1861.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Lilley G. G., Stewart D. J., Kortt A. A. Amino acid and DNA sequences of an extracellular basic protease of Dichelobacter nodosus show that it is a member of the subtilisin family of proteases. Eur J Biochem. 1992 Nov 15;210(1):13–21. doi: 10.1111/j.1432-1033.1992.tb17385.x. [DOI] [PubMed] [Google Scholar]
  7. Paradis F. W., Warren R. A., Kilburn D. G., Miller R. C., Jr The expression of Cellulomonas fimi cellulase genes in Brevibacterium lactofermentum. Gene. 1987;61(2):199–206. doi: 10.1016/0378-1119(87)90114-4. [DOI] [PubMed] [Google Scholar]
  8. Park S. S., Wong S. L., Wang L. F., Doi R. H. Bacillus subtilis subtilisin gene (aprE) is expressed from a sigma A (sigma 43) promoter in vitro and in vivo. J Bacteriol. 1989 May;171(5):2657–2665. doi: 10.1128/jb.171.5.2657-2665.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Radford A. J., Hodgson A. L. Construction and characterization of a Mycobacterium-Escherichia coli shuttle vector. Plasmid. 1991 Mar;25(2):149–153. doi: 10.1016/0147-619x(91)90029-v. [DOI] [PubMed] [Google Scholar]
  10. Smith M. D., Flickinger J. L., Lineberger D. W., Schmidt B. Protoplast transformation in coryneform bacteria and introduction of an alpha-amylase gene from Bacillus amyloliquefaciens into Brevibacterium lactofermentum. Appl Environ Microbiol. 1986 Mar;51(3):634–639. doi: 10.1128/aem.51.3.634-639.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Wang L. F., Doi R. H. Developmental expression of three proteins from the first gene of the RNA polymerase sigma 43 operon of Bacillus subtilis. J Bacteriol. 1987 Sep;169(9):4190–4195. doi: 10.1128/jb.169.9.4190-4195.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Wang L. F., Kortt A. A., Stewart D. J. Use of a gram- signal peptide for protein secretion by gram+ hosts: basic protease of Dichelobacter nodosus is produced and secreted by Bacillus subtilis. Gene. 1993 Sep 6;131(1):97–102. doi: 10.1016/0378-1119(93)90675-s. [DOI] [PubMed] [Google Scholar]
  13. Wells J. A., Ferrari E., Henner D. J., Estell D. A., Chen E. Y. Cloning, sequencing, and secretion of Bacillus amyloliquefaciens subtilisin in Bacillus subtilis. Nucleic Acids Res. 1983 Nov 25;11(22):7911–7925. doi: 10.1093/nar/11.22.7911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Wu Z. R., Qi B. J., Jiao R. Q., Chen F. D., Wang L. F. Development of a novel Bacillus subtilis cloning system employing its neutral protease as screen marker. Gene. 1991 Sep 30;106(1):103–107. doi: 10.1016/0378-1119(91)90572-s. [DOI] [PubMed] [Google Scholar]
  15. Yoshihama M., Higashiro K., Rao E. A., Akedo M., Shanabruch W. G., Follettie M. T., Walker G. C., Sinskey A. J. Cloning vector system for Corynebacterium glutamicum. J Bacteriol. 1985 May;162(2):591–597. doi: 10.1128/jb.162.2.591-597.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES