Abstract
Strain S2 is a lecithin (or phosphatidylcholine)-solubilizing bacterium, which was isolated from the rice rhizosphere in rural areas of Beijing, China. On the basis of a polyphasic study involving phenotypic tests, physiological and biochemical tests, 16S rDNA sequence analysis, G+C content determination and DNA-DNA hybridizations analysis, strain S2 was identified as Pseudomonas alcaligenes. P. alcaligenes S2 was mutagenized with Tn5 and four mutants showing decreased or increased solubilizing ability of lecithin were isolated based on the halo size around colonies on the solid plate supplemented with egg yolk. To characterize the genes of P. alcaligenes S2 involved in solubilization of lecithin, the EcoR I fragments of the chromosomes from the four mutant strains carrying a single transposon were cloned, and the DNA sequences flanking the Tn5 were determined. The Tn5 insertion sites in the mutants M808, M1329 and M1400, showing decreased solubilizing ability of lecithin, were found to be located in the xcpS, xcpX and xcpW, respectively, whose products XcpS, XcpX and XcpW were the components of type II secretion pathway. Complementation of xcpS, xcpX and xcpW could restore the corresponding mutants M808, M1329 and M1400 to solubilize lecithin. The data suggested that mutation in one of these xcp genes would lead to the absence of mature lecithinase secretion into the extracellular medium. The data also indicated that the secretion of lecithin-hydrolyzing enzyme of P. alcaligeneswas via type II secretion pathway. In the mutant M20 showing increasing lecithin-hydrolyzing activity, the interrupted gene showed 86% identity with chpA of Pseudomonas aeruginosa PAO1, whose product plays an important role in controlling twitching motility of the bacterial cells.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Rodriguez, H., Fraga, R., Phosphate solubilizing bacteria and their role in plant growth promotion, Biotechnol. Adv., 1999, 17: 319–339.
Turner, B. L., Cade-Menum, B. J., Westermann, D. T., Organic phosphorus Composition and potential bioavailability in semi-arid arable soils of the western United States, Soil Sci. Soc. AM. J., 2003, 67: 1168–1179.
Abd-Alla, M. H., Use of organic phosphorus by Rhizobium leguminosarum biovar. viceae phosphatases, Biol. Fertil. Soils, 1994, 18: 216–218.
Thaller, M. C., Berlutti, F., Schippa, S. et al., Heterogeneous patterns of acid phosphatases containing low-molecular-mass polipeptides in members of the family Enterobacteriaceae, Int. J. Syst. Bacteriol., 1995, 4: 255–261.
Skrary, F. A., Cameron, D. C., Purification and characterization of a Bacillus licheniformis phosphatase specific for D-alpha-glycerphosphate, Arch. Biochem. Biophys., 1998, 349: 27–35.
Richardson, A. E., Hadobas, P. A., Soil isolates of Pseudomonas spp. that utilize inositol phosphates, Can. J. Microbial., 1997, 43: 509–516.
Thaler, J. O., Duvic, B., Givaudan, A., Isolation and entomotoxic properties of the Xenorhabdus nematophilus F1 lecithinase, Appl. Environ. Microbiol., 1998, 64: 2367–2373.
Wolk, C. P., Cai, Y., Jean-michel, P., Use of a transposon with luciferase as a reporter to identify environmentally responsive genes in a cyanobacterium, Proc. Nat. Acad. Sci. USA, 1991, 88: 5355–5359.
Staskawicz, B. D., Dahlbeck, N. K., Napoli, C., Molecular characterization of cloned a virulence genes from race 0 and race 1 of Pseudomonas syringae pv. Glycinea, J. Bacteriol., 1987, 169: 5789–5794.
Buchanan, R. E., Gibbons, N. E., Genus II, Pseudomonas, in Bergey’s Manual of Systematic Bacteriology (eds. Krieg, N. R., Holt, J. G.), Baltimore: Williams Wilkins, 1984, 1: 141–198.
Brenner, D. J., McWhorter, A. C., Knutson, J. K., et al., Escherichia vulneris: A species of Enterobacteriaceae associated with human wounds, J. Clin. Microbiol., 1982, 15, 133–1140.
Crosa, J. H., Brenner, D. J., Falkow, S., Use of a single-strand specific nuclease for analysis of bacterial and plasmid deoxyribonucleic acid homoduplex and heteroduplex, J. Bacteriol., 1973, 115: 904–911.
Marmur, J., Doty, P., Determination of the base composition of deoxyribonucleic acid from its thermal denaturation temperature, J. Mol. Biol., 1962, 5: 109–118.
Kim, K., Lee, S., Lee, K. et al., Isolation and characterization of toluene-sensitive mutants from the toluene-resistant bacterium Pseudomonas putida GM73, J. Bacteriol., 1998, 180: 3692–3696.
Sambrook, J., Russell, D. W., Molecular cloning: A laboratory manual, 3rd ed, New York: Cold Spring Harbor Laboratory Press, 2001.
Olsen, S. R., Sommers, L. E., Phosphorus, in Methods of Soil Analysis, Part 2: Chemical and Microbiological Properties (eds. Page, A. L., Miller, R. H., Keeney, D. R.), 2nd ed, Madison: ASA-SSSA, 1982, 9 (2): 403–430.
Wretlind, B., Pavlovskis, O. R., Genetic mapping and characterization of Pseudomonas aeruginosa mutants defective in the formation of extracellular proteins, J. Bacteriol., 1984, 158: 801–808.
Ball, G., Durand, E., Lazdunski, A., A novel type II secretion system in Pseudomonas aeruginosa, Mol. Microbiol., 2002, 43: 475–485.
Filloux, A., Michel, G., Bally, M., GSP-dependent protein secretion in Gram-negative bacteria: The Xcp machinery of Pseudomonas aeruginosa, FEMS Microbiol. Rev., 1998, 22: 177–198.
Thanassi, D. G., Hultgren, S. J., Multiple pathways allow protein secretion across the bacterial outer membrane, Curr. Opin. Cell Biol., 2000, 12: 420–430.
Pugsley, A. P., The complete general secretory pathway in Gram negative bacteria, Microbiol. Rev., 1993, 57: 50–108.
Duong, F., Soscia, A., Lazdunski, A., et al., The Pseudomonas fluorescens lipase has a C-terminal secretion signal and is secreted by a three-component bacterial ABC-exporter system, Mol. Microbiol., 1994, 11: 1117–1126.
Bitter, W., Koster, M., Tommassen, J., Formation of oligomeric rings by XcpQ and PhilQ, which are involved in protein transport across the outer membrane of Pseudomonas aeruginosa, Mol. Microbiol., 1998, 27: 209–219.
Duong, F., Bonnet, E., Geli, V., The AprX protein of Pseudomonas aeruginosa: A new substrate for the Apr type I secretion system, Gene, 2001, 262: 147–153.
Frank, D. W., The exoenzyme S regulon of Pseudomonas aeruginosa, Mol. Microbiol., 1997, 26: 621–629.
Akatsuka, H., Kawai, E., Omori, K., The three genes lipB, lipC, and lipD involved in the extracellular secretion of the Serratia marcescens lipase which lacks an N-terminal signal peptide, J. Bacteriol., 1995, 177: 6381–6389.
Kawai, E., Idei, A., Kumura, H. et al., The ABC-exporter genes involved in the lipase secretion are clustered with the genes for lipase, alkaline protease, and serine protease homologues in Pseudomonas fluorescens, Biochim. Biophys. acta, 1999, 1446: 377–382.
Whitchurch, C. B., Leech, A. J., Young, M. D. et al., Characterization of a complex chemosensory signal transduction system which controls twitching motility in Pseudomonas aeruginosa, Mol. Microbiol., 2004, 52: 873–893.
Cynthia, B. W., Andrew, J. L., Michael, D. Y., Characterization of a complex chemosensory signal transduction system which controls twitching motility in Pseudomonas aeruginosa, Mol. Microbiol., 2004, 52: 873–893.
Author information
Authors and Affiliations
Corresponding author
About this article
Cite this article
Lü, J., Li, F., Chen, S. et al. The secretion of lecithinase of Pseudomonas alcaligenes S2 was via type II secretion pathway. Chin.Sci.Bull. 50, 1731–1736 (2005). https://doi.org/10.1360/982005-548
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1360/982005-548