This Article
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Franklin, M J
Right arrow Articles by Ohman, D E
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Franklin, M J
Right arrow Articles by Ohman, D E

 Previous Article  |  Next Article 

J Bacteriol. 1993 August; 175(16): 5057-5065

research-article

Identification of algF in the alginate biosynthetic gene cluster of Pseudomonas aeruginosa which is required for alginate acetylation.

M J Franklin and D E Ohman

Department of Microbiology and Immunology, University of Tennessee, Memphis.

ABSTRACT

Mucoid strains of Pseudomonas aeruginosa produce a high-molecular-weight exopolysaccharide called alginate that is modified by the addition of O-acetyl groups. To better understand the acetylation process, a gene involved in alginate acetylation called algF was identified in this study. We hypothesized that a gene involved in alginate acetylation would be located within the alginate biosynthetic gene cluster at 34 min on the P. aeruginosa chromosome. To isolate algF mutants, a procedure for localized mutagenesis was developed to introduce random chemical mutations into the P. aeruginosa alginate biosynthetic operon on the chromosome. For this, a DNA fragment containing the alginate biosynthetic operon and adjacent argF gene in a gene replacement cosmid vector was utilized. The plasmid was packaged in vivo into lambda phage particles, mutagenized in vitro with hydroxylamine, transduced into Escherichia coli, and mobilized to an argF auxotroph of P. aeruginosa FRD. Arg+ recombinants coinherited the mutagenized alginate gene cluster and were screened for defects in alginate acetylation by testing for increased sensitivity to an alginate lyase produced by Klebsiella aerogenes. Alginates from recombinants which showed increased sensitivity to alginate lyase were tested for acetylation by a colorimetric assay and infrared spectroscopy. Two algF mutants that produced alginates reduced more than sixfold in acetyl groups were obtained. The acetylation defect was complemented in trans by a 3.8-kb XbaI-BamHI fragment from the alginate gene cluster when placed in the correct orientation under a trc promoter. By a merodiploid analysis, the algF gene was further mapped to a region directly upstream of algA by examining the polar effect of Tn501 insertions. By gene replacement, DNA with a Tn501 insertion directly upstream of algA was recombined with the chromosome of mucoid strain FRD1. The resulting strain, FRD1003, was nonmucoid because of the polar effect of the transposon on the downstream algA gene. By providing algA in trans under the tac promoter, FRD1003 produced nonacetylated alginate, indicating that the transposon was within or just upstream of algF. These results demonstrated that algF, a gene involved in alginate acetylation, is located directly upstream of algA.

J Bacteriol. 1993 August; 175(16): 5057-5065




This article has been cited by other articles:

  • Son, M. S., Matthews, W. J. Jr., Kang, Y., Nguyen, D. T., Hoang, T. T. (2007). In Vivo Evidence of Pseudomonas aeruginosa Nutrient Acquisition and Pathogenesis in the Lungs of Cystic Fibrosis Patients. Infect. Immun. 75: 5313-5324 [Abstract] [Full Text]  
  • Remminghorst, U., Rehm, B. H. A. (2006). In Vitro Alginate Polymerization and the Functional Role of Alg8 in Alginate Production by Pseudomonas aeruginosa. Appl. Environ. Microbiol. 72: 298-305 [Abstract] [Full Text]  
  • Jain, S., Ohman, D. E. (2005). Role of an Alginate Lyase for Alginate Transport in Mucoid Pseudomonas aeruginosa. Infect. Immun. 73: 6429-6436 [Abstract] [Full Text]  
  • Douthit, S. A., Dlakic, M., Ohman, D. E., Franklin, M. J. (2005). Epimerase Active Domain of Pseudomonas aeruginosa AlgG, a Protein That Contains a Right-Handed {beta}-Helix. J. Bacteriol. 187: 4573-4583 [Abstract] [Full Text]  
  • Albrecht, M. T., Schiller, N. L. (2005). Alginate Lyase (AlgL) Activity Is Required for Alginate Biosynthesis in Pseudomonas aeruginosa. J. Bacteriol. 187: 3869-3872 [Abstract] [Full Text]  
  • Robles-Price, A., Wong, T. Y., Sletta, H., Valla, S., Schiller, N. L. (2004). AlgX Is a Periplasmic Protein Required for Alginate Biosynthesis in Pseudomonas aeruginosa. J. Bacteriol. 186: 7369-7377 [Abstract] [Full Text]  
  • Franklin, M. J., Douthit, S. A., McClure, M. A. (2004). Evidence that the algI/algJ Gene Cassette, Required for O Acetylation of Pseudomonas aeruginosa Alginate, Evolved by Lateral Gene Transfer. J. Bacteriol. 186: 4759-4773 [Abstract] [Full Text]  
  • Franklin, M. J., Ohman, D. E. (2002). Mutant Analysis and Cellular Localization of the AlgI, AlgJ, and AlgF Proteins Required for O Acetylation of Alginate in Pseudomonas aeruginosa. J. Bacteriol. 184: 3000-3007 [Abstract] [Full Text]  
  • Pier, G. B., Coleman, F., Grout, M., Franklin, M., Ohman, D. E. (2001). Role of Alginate O Acetylation in Resistance of Mucoid Pseudomonas aeruginosa to Opsonic Phagocytosis. Infect. Immun. 69: 1895-1901 [Abstract] [Full Text]  
  • Nivens, D. E., Ohman, D. E., Williams, J., Franklin, M. J. (2001). Role of Alginate and Its O Acetylation in Formation of Pseudomonas aeruginosa Microcolonies and Biofilms. J. Bacteriol. 183: 1047-1057 [Abstract] [Full Text]  
  • Bolduc, G. R., Bouchet, V., Jiang, R.-Z., Geisselsoder, J., Truong-Bolduc, Q. C., Rice, P. A., Pelton, S. I., Goldstein, R. (2000). Variability of Outer Membrane Protein P1 and Its Evaluation as a Vaccine Candidate against Experimental Otitis Media due to Nontypeable Haemophilus influenzae: an Unambiguous, Multifaceted Approach. Infect. Immun. 68: 4505-4517 [Abstract] [Full Text]  
  • Fakhr, M. K., Peñaloza-Vázquez, A., Chakrabarty, A. M., Bender, C. L. (1999). Regulation of Alginate Biosynthesis in Pseudomonas syringae pv. syringae. J. Bacteriol. 181: 3478-3485 [Abstract] [Full Text]  
  • Hatch, R. A., Schiller, N. L. (1998). Alginate Lyase Promotes Diffusion of Aminoglycosides through the Extracellular Polysaccharide of Mucoid Pseudomonas aeruginosa. Antimicrob. Agents Chemother. 42: 974-977 [Abstract] [Full Text]  
  • Jain, S., Ohman, D. E. (1998). Deletion of algK in Mucoid Pseudomonas aeruginosa Blocks Alginate Polymer Formation and Results in Uronic Acid Secretion. J. Bacteriol. 180: 634-641 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»