Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1982 Dec;79(24):7881–7885. doi: 10.1073/pnas.79.24.7881

Genetic and biochemical analysis of gonococcal IgA1 protease: cloning in Escherichia coli and construction of mutants of gonococci that fail to produce the activity.

J M Koomey, R E Gill, S Falkow
PMCID: PMC347453  PMID: 6818556

Abstract

The biological significance of bacterial extracellular proteases that specifically cleave human IgA1 is unknown. We have prepared a gene bank of gonococcal chromosomal DNA in Escherichia coli K-12 using a cosmid cloning system. Among these clones, we have identified and characterized an E. coli strain that elaborates an extracellular endopeptidase that is indistinguishable from gonococcal IgA1 protease in its substrate specificity and action on human IgA1. Analysis of recombinant plasmids and examination of plasmid-specific peptides in minicells have shown that the IgA1 protease activity in E. coli is associated with expression of a Mr 140,000 peptide. We have isolated IgA1 protease-deficient mutants of Neisseria gonorrhoeae by reintroduction of physically defined deletions of the cloned gene into the gonococcal chromosome by transformation.

Full text

PDF
7881

Images in this article

7882Image
on p.7882
7883Image
on p.7883
7884Image
on p.7884
7885Image
on p.7885

Selected References

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

  1. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blake M. S., Swanson J. Studies on gonococcus infection. XVI. Purification of Neisseria gonorrhoeae immunoglobulin A1 protease. Infect Immun. 1978 Nov;22(2):350–358. doi: 10.1128/iai.22.2.350-358.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  4. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  5. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dougan G., Sherratt D. The transposon Tn1 as a probe for studying ColE1 structure and function. Mol Gen Genet. 1977 Mar 7;151(2):151–160. doi: 10.1007/BF00338689. [DOI] [PubMed] [Google Scholar]
  7. Elwell L. P., Roberts M., Mayer L. W., Falkow S. Plasmid-mediated beta-lactamase production in Neisseria gonorrhoeae. Antimicrob Agents Chemother. 1977 Mar;11(3):528–533. doi: 10.1128/aac.11.3.528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Garten W., Hindennach I., Henning U. The major proteins of the Escherichia coli outer cell envelope membrane. Characterization of proteins II* and III, comparison of all proteins. Eur J Biochem. 1975 Nov 1;59(1):215–221. doi: 10.1111/j.1432-1033.1975.tb02444.x. [DOI] [PubMed] [Google Scholar]
  9. Genco R. J., Plaut A. G., Moellering R. C., Jr Evaluation of human oral organisms and pathogenic Streptococcus for production of IgA protease. J Infect Dis. 1975 May;131 (Suppl):S17–S21. doi: 10.1093/infdis/131.supplement.s17. [DOI] [PubMed] [Google Scholar]
  10. Gill R. E., Heffron F., Falkow S. Identification of the protein encoded by the transposable element Tn3 which is required for its transposition. Nature. 1979 Dec 20;282(5741):797–801. doi: 10.1038/282797a0. [DOI] [PubMed] [Google Scholar]
  11. Hull R. A., Gill R. E., Hsu P., Minshew B. H., Falkow S. Construction and expression of recombinant plasmids encoding type 1 or D-mannose-resistant pili from a urinary tract infection Escherichia coli isolate. Infect Immun. 1981 Sep;33(3):933–938. doi: 10.1128/iai.33.3.933-938.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kilian M. Degradation of immunoglobulins A2, A2, and G by suspected principal periodontal pathogens. Infect Immun. 1981 Dec;34(3):757–765. doi: 10.1128/iai.34.3.757-765.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kilian M., Mestecky J., Kulhavy R., Tomana M., Butler W. T. IgA1 proteases from Haemophilus influenzae, Streptococcus pneumoniae, Neisseria meningitidis, and Streptococcus sanguis: comparative immunochemical studies. J Immunol. 1980 Jun;124(6):2596–2600. [PubMed] [Google Scholar]
  14. Kilian M., Mestecky J., Schrohenloher R. E. Pathogenic species of the genus Haemophilus and Streptococcus pneumoniae produce immunoglobulin A1 protease. Infect Immun. 1979 Oct;26(1):143–149. doi: 10.1128/iai.26.1.143-149.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kornfeld S. J., Plaut A. G. Secretory immunity and the bacterial IgA proteases. Rev Infect Dis. 1981 May-Jun;3(3):521–534. doi: 10.1093/clinids/3.3.521. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Male C. J. Immunoglobulin A1 protease production by Haemophilus influenzae and Streptococcus pneumoniae. Infect Immun. 1979 Oct;26(1):254–261. doi: 10.1128/iai.26.1.254-261.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Mathis L. S., Scocca J. J. Haemophilus influenzae and Neisseria gonorrhoeae recognize different specificity determinants in the DNA uptake step of genetic transformation. J Gen Microbiol. 1982 May;128(5):1159–1161. doi: 10.1099/00221287-128-5-1159. [DOI] [PubMed] [Google Scholar]
  19. Mayer L. W., Holmes K. K., Falkow S. Characterization of plasmid deoxyribonucleic acid from Neisseria gonorrhoeae. Infect Immun. 1974 Oct;10(4):712–717. doi: 10.1128/iai.10.4.712-717.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mulks M. H., Kornfeld S. J., Plaut A. G. Specific proteolysis of human IgA by Streptococcus pneumoniae and Haemophilus influenzae. J Infect Dis. 1980 Apr;141(4):450–456. doi: 10.1093/infdis/141.4.450. [DOI] [PubMed] [Google Scholar]
  21. Plaut A. G., Gilbert J. V., Artenstein M. S., Capra J. D. Neisseria gonorrhoeae and neisseria meningitidis: extracellular enzyme cleaves human immunoglobulin A. Science. 1975 Dec 12;190(4219):1103–1105. doi: 10.1126/science.810892. [DOI] [PubMed] [Google Scholar]
  22. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  23. Roberts M., Elwell L. P., Falkow S. Molecular characterization of two beta-lactamase-specifying plasmids isolated from Neisseria gonorrhoeae. J Bacteriol. 1977 Aug;131(2):557–563. doi: 10.1128/jb.131.2.557-563.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Roozen K. J., Fenwick R. G., Jr, Curtiss R., 3rd Synthesis of ribonucleic acid and protein in plasmid-containing minicells of Escherichia coli K-12. J Bacteriol. 1971 Jul;107(1):21–33. doi: 10.1128/jb.107.1.21-33.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  26. Sparling P. F. Genetic transformation of Neisseria gonorrhoeae to streptomycin resistance. J Bacteriol. 1966 Nov;92(5):1364–1371. doi: 10.1128/jb.92.5.1364-1371.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sternberg N., Tiemeier D., Enquist L. In vitro packaging of a lambda Dam vector containing EcoRI DNA fragments of Escherichia coli and phage P1. Gene. 1977 May;1(3-4):255–280. doi: 10.1016/0378-1119(77)90049-x. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES