Construction of isogenic urease-negative mutants of Helicobacter pylori by allelic exchange - PubMed (original) (raw)
Construction of isogenic urease-negative mutants of Helicobacter pylori by allelic exchange
R L Ferrero et al. J Bacteriol. 1992 Jul.
Abstract
Isogenic urease-negative mutants of Helicobacter pylori were constructed by allelic replacement. A region of cloned H. pylori DNA containing the structural urease genes (ureA and ureB) was disrupted by insertion of a mini-Tn3-Km transposon. Electrotransformation of H. pylori cells with kanamycin-ureB-disrupted derivative plasmids resulted in isolation of kanamycin-resistant H. pylori transformants. Competence for electrotransformation appeared to be restricted to certain wild-type H. pylori isolates; only 1 isolate (of 10 tested) was consistently transformed. Two of the kanamycin-resistant H. pylori transformants were further studied and shown to be urease negative. Southern hybridization analyses demonstrated that the urease-negative mutants had been constructed by allelic exchange involving simultaneous replacement of the ureB gene with the kanamycin-ureB-disrupted copy and loss of the vector. Immunoblot studies of whole-cell extracts of the isogenic ureB mutants with anti-H. pylori sera indicated the absence of a polypeptide with an apparent molecular mass of 61 kDa; thus, the mutants no longer synthesized the UreB product. Generation of stable, genetically engineered urease mutants of H. pylori will be useful for addressing the role of urease in the pathogenesis of H. pylori infection.
Similar articles
- Purification of recombinant Helicobacter pylori urease apoenzyme encoded by ureA and ureB.
Hu LT, Foxall PA, Russell R, Mobley HL. Hu LT, et al. Infect Immun. 1992 Jul;60(7):2657-66. doi: 10.1128/iai.60.7.2657-2666.1992. Infect Immun. 1992. PMID: 1612735 Free PMC article. - Expression of Helicobacter pylori urease genes in Escherichia coli grown under nitrogen-limiting conditions.
Cussac V, Ferrero RL, Labigne A. Cussac V, et al. J Bacteriol. 1992 Apr;174(8):2466-73. doi: 10.1128/jb.174.8.2466-2473.1992. J Bacteriol. 1992. PMID: 1313413 Free PMC article. - Construction of a Helicobacter pylori-Escherichia coli shuttle vector for gene transfer in Helicobacter pylori.
Lee WK, An YS, Kim KH, Kim SH, Song JY, Ryu BD, Choi YJ, Yoon YH, Baik SC, Rhee KH, Cho MJ. Lee WK, et al. Appl Environ Microbiol. 1997 Dec;63(12):4866-71. doi: 10.1128/aem.63.12.4866-4871.1997. Appl Environ Microbiol. 1997. PMID: 9406406 Free PMC article. - Direct random insertion mutagenesis of Helicobacter pylori.
de Jonge R, Bakker D, van Vliet AH, Kuipers EJ, Vandenbroucke-Grauls CM, Kusters JG. de Jonge R, et al. J Microbiol Methods. 2003 Jan;52(1):93-100. doi: 10.1016/s0167-7012(02)00136-7. J Microbiol Methods. 2003. PMID: 12401231 - Shuttle cloning and nucleotide sequences of Helicobacter pylori genes responsible for urease activity.
Labigne A, Cussac V, Courcoux P. Labigne A, et al. J Bacteriol. 1991 Mar;173(6):1920-31. doi: 10.1128/jb.173.6.1920-1931.1991. J Bacteriol. 1991. PMID: 2001995 Free PMC article.
Cited by
- Campylobacter jejuni extracellular vesicles harboring cytolethal distending toxin bind host cell glycans and induce cell cycle arrest in host cells.
Le LHM, Elgamoudi B, Colon N, Cramond A, Poly F, Ying L, Korolik V, Ferrero RL. Le LHM, et al. Microbiol Spectr. 2024 Mar 5;12(3):e0323223. doi: 10.1128/spectrum.03232-23. Epub 2024 Feb 6. Microbiol Spectr. 2024. PMID: 38319111 Free PMC article. - Trimer stability of Helicobacter pylori HtrA is regulated by a natural mutation in the protease domain.
Zarzecka U, Tegtmeyer N, Sticht H, Backert S. Zarzecka U, et al. Med Microbiol Immunol. 2023 Jun;212(3):241-252. doi: 10.1007/s00430-023-00766-9. Epub 2023 May 14. Med Microbiol Immunol. 2023. PMID: 37183214 Free PMC article. - Helicobacter pylori Modulates Heptose Metabolite Biosynthesis and Heptose-Dependent Innate Immune Host Cell Activation by Multiple Mechanisms.
Hauke M, Metz F, Rapp J, Faass L, Bats SH, Radziej S, Link H, Eisenreich W, Josenhans C. Hauke M, et al. Microbiol Spectr. 2023 Jun 15;11(3):e0313222. doi: 10.1128/spectrum.03132-22. Epub 2023 Apr 27. Microbiol Spectr. 2023. PMID: 37129481 Free PMC article. - Identification of Antimotilins, Novel Inhibitors of Helicobacter pylori Flagellar Motility That Inhibit Stomach Colonization in a Mouse Model.
Suerbaum S, Coombs N, Patel L, Pscheniza D, Rox K, Falk C, Gruber AD, Kershaw O, Chhatwal P, Brönstrup M, Bilitewski U, Josenhans C. Suerbaum S, et al. mBio. 2022 Apr 26;13(2):e0375521. doi: 10.1128/mbio.03755-21. Epub 2022 Mar 1. mBio. 2022. PMID: 35227071 Free PMC article. - Helicobacter pylori virulence factors: relationship between genetic variability and phylogeographic origin.
Rodriguez AM, Urrea DA, Prada CF. Rodriguez AM, et al. PeerJ. 2021 Nov 26;9:e12272. doi: 10.7717/peerj.12272. eCollection 2021. PeerJ. 2021. PMID: 34900406 Free PMC article.
References
- Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350-4 - PubMed
- Nature. 1970 Aug 15;227(5259):680-5 - PubMed
- J Bacteriol. 1992 Apr;174(8):2466-73 - PubMed
- Res Microbiol. 1992 Jan;143(1):15-26 - PubMed
- N Engl J Med. 1991 Oct 17;325(16):1127-31 - PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources