Characterization of the novel nisin-sucrose conjugative transposon Tn5276 and its insertion in Lactococcus lactis
Tóm tắt
A novel, chromosomally located conjugative transposon in Lactococcus lactis, Tn5276, was identified and characterized. It encodes the production of and immunity to nisin, a lanthionine-containing peptide with antimicrobial activity, and the capacity to utilize sucrose via a phosphotransferase system. Conjugal transfer of Tn5276 was demonstrated from L. lactis NIZO R5 to different L. lactis strains and a recombination-deficient mutant. The integration of Tn5276 into the plasmid-free strain MG1614 was analyzed by using probes based on the gene for the nisin precursor (nisA) and the gene for sucrose-6-phosphate hydrolase (sacA). The transposon inserted at various locations in the MG1614 chromosome and showed a preference for orientation-specific insertion into a single target site (designated site 1). By using restriction mapping in combination with field inversion gel electrophoresis and DNA cloning of various parts of the element including its left and right ends, a physical map of the 70-kb Tn5276 was constructed, and the nisA and sacA genes were located. The nucleotide sequences of Tn5276 junctions in donor strain NIZO R5 and in site 1 of an MG1614-derived transconjugant were determined and compared with that of site 1 in recipient strain MG1614. The results show that the A + T-rich ends of Tn5276 are flanked by a direct hexanucleotide repeat in both the donor and the transconjugant but that the element does not contain a clear inverted repeat.
Từ khóa
Tài liệu tham khảo
Anderson , D. G. , and L. L. McKay . 1983 . Isolation of a recombination-deficient mutant of Streptococcus lactis ML3 . J. Bacteriol. 155 : 930 - 932 .
Buchman , W. B. , S. Banerjee , and J. R. Hansen . 1988 . Structure, expression and evolution of a gene encoding the precursor of nisin, a small protein antibiotic . J. Biol. Chem. 263 : 16260 - 16266 .
Caillaud , F. , and P. Courvalin . 1987 . Nucleotide sequence of the ends of the conjugative shuttle transposon TnlS45 . Mol. Gen. Genet. 209 : 110 - 115 .
Campbell , A. , D. E. Berg , D. Botstein , E. M. Lederberg , R. P. Novick , P. Starlinger , and W. Szybalski . 1979 . Nomenclature of transposable elements in prokaryotes . Gene 5 : 197 - 206 .
Caparon , M. G. , and J. R. Scott . 1989 . Excision and insertion of the conjugative transposon Tn 916 involves a novel recombination mechanism . Cell 59 : 1027 - 1034 .
Casadaban , M. J. , and S. N. Cohen . 1980 . Analysis of gene control signals by DNA fusion and cloning in E. coli . J. Mol. Biol. 143 : 179 - 207 .
Clewell , D. B. , S. E. Flannagan , Y. Ike , J. M. Jones , and C. Gawron-Burke . 1988 . Sequence analysis of termini of conjugative transposon Tn 916 . J. Bacteriol. 170 : 3046 - 3052 .
Clewell , D. B. , and C. Gawron-Burke . 1986 . Conjugative transposons and the dissemination of antibiotic resistance in streptococci. Annu . Rev. Microbiol. 40 : 635 - 659 .
Dodd , H. M. , N. Horn , and M. J. Gasson . 1990 . Analysis of the genetic determinant for production of the peptide antibiotic nisin . J. Gen. Microbiol. 136 : 555 - 566 .
Donkersloot , J. A. , and J. Thompson . 1990 . Simultaneous loss of N5-(carboxyethyl)ornithine synthase, nisin production, and sucrose-fermenting ability by Lactococcus lactis Kl . J. Bacteriol. 172 : 4122 - 4126 .
Elliker , P. R. , A. Anderson , and G. H. Hannessen . 1956 . An agar culture medium for lactic streptococci and lactobacilli . J. Dairy Sci. 39 : 1611 - 1612 .
Fitzgerald , G. F. , and M. J. Gasson . 1988 . In vivo gene transfer systems and transposons . Biochimie 70 : 489 - 502 .
Frischauf , A. M. , M. Lehrach , A. Poustka , and N. Murray . 1983 . Lambda replacement vectors carrying polylinker sequences . J. Mol. Biol. 170 : 827 - 842 .
Galesloot , T. E. , and J. W. Pette . 1957 . The formation of normal eyes in Edam cheese made by means of nisin-producing starters . Neth. Milk Dairy J. 11 : 144 - 151 .
Gasson , M. J. 1983 . Plasmid complements of Streptococcus lactis NCDO 712 and other lactic streptococci after protoplastinduced curing . J. Bacteriol. 154 : 1 - 9 .
Gasson , M. J. 1984 . Transfer of sucrose fermenting ability, nisin resistance and nisin production into Streptococcus lactis 712 . FEMS Microbiol. Lett. 21 : 7 - 10 .
Gasson , M. J. , and F. L. Davies . 1980 . Conjugal transfer of the drug resistance plasmid pAMP in the lactic streptococci . FEMS Microbiol. Lett. 7 : 51 - 53 .
Gawron-Burke , C. , and D. B. Clewell . 1982 . A transposon in Streptococcus faecalis with fertility properties . Nature (London) 300 : 281 - 284 .
Gibson T. J. 1984. Studies on the Eppstein-Barr genome. Ph.D. thesis Cambridge University Cambridge.
Gonzalez , C. F. , and B. S. Kunka . 1985 . Transfer of sucrose fermenting ability and nisin production phenotype among lactic streptococci . Appl. Environ. Microbiol. 49 : 627 - 633 .
Hattori , M. , and Y. Sakaki . 1986 . Dideoxy sequencing using denatured plasmid templates . Anal. Biochem. 152 : 232 .
Hill , C. , C. Daly , and G. F. Fitzgerald . 1985 . Conjugative transfer of the transposon Tn 919 to lactic acid bacteria . FEMS Microbiol. Lett. 30 : 115 - 119 .
Hill , C. , C. Daly , and G. F. Fitzgerald . 1987 . Development of high-frequency delivery system for transposon Tn 919 in lactic streptococci: random insertion in Streptococcus lactis subsp. diacetylactis 18-16 . Appl. Environ. Microbiol. 53 : 74 - 78 .
Horne , N. , H. M. Dodd , and M. J. Gasson . 1991 . Nisin biosynthesis genes are encoded by a novel conjugative transposon . Mol. Gen. Genet. 228 : 129 - 135 .
Inamine , J. M. , and V. Burdett . 1985 . Structural organization of a 67-kilobase streptococcal conjugative element mediating multiple antibiotic resistance . J. Bacteriol. 161 : 620 - 626 .
Kaletta , C. , and K. -D. Entian . 1989 . Nisin, a peptide antibiotic: cloning and sequencing of the nisA gene and post-translational processing of its peptide product . J. Bacteriol. 171 : 1597 - 1601 .
Krolewski , J. J. , E. Murphy , R. P. Novick , and M. G. Rush . 1981 . Site-specificity of the chromosomal insertion of Staphylococcus aureus transposon TnS54 . J. Mol. Biol. 152 : 19 - 33 .
Lacks S. A. 1988. Mechanisms of genetic recombination in gram-positive bacteria p. 43-86. In R. Kucherlapati and G. R. Smith (ed.) Genetic recombination. American Society for Microbiology Washington D.C.
Leblanc D. J. V. L. Crow and L. N. Lee. 1980. Plasmid mediated carbohydrate catabolic enzymes among strains of Streptococcus lactis p. 31-41. In C. Studdard and K. R. Rozee (ed.) Plasmids and transposons: environmental effects and maintenance mechanisms. Academic Press Inc. New York.
Le Bouguenec , C. , G. de Cespedes , and T. Horaud . 1988 . Molecular analysis of a composite chromosomal conjugative element (Tn 3701) of Streptococcus pyogenes . J. Bacteriol. 170 : 3930 - 3936 .
Lederberg , E. M. 1987 . Plasmid reference centre registry of transposon (Tn) and insertion sequence (IS) allocations through December, 1986 . Gene 51 : 115 - 118 .
Mulders , J. W. M. , I. J. Boerrigter , H. S. Rollema , R. J. Siezen , and W. M. de Vos . 1991 . Identification and characterization of the lantibiotic nisin Z, a natural nisin variant . Eur. J. Biochem. 201 : 581 - 584 .
Murphy E. 1989. Transposable elements in gram-positive bacteria p. 269-288. In D. E. Berg and M. M. Howe (ed.) Mobile DNA. American Society for Microbiology Washington D.C.
Murphy , E. , L. Huwyler , and M. C. F. Bastos . 1985 . Transposon TnS54: complete nucleotide sequence and isolation of transposition-defective and antibiotic-sensitive mutants . EMBO J. 4 : 3357 - 3365 .
Murphy , M. C. , J. L. Steele , C. Daly , and L. L. McKay . 1988 . Concomitant conjugal transfer of reduced bacteriophage sensitivity mechanisms with lactose- and sucrose-fermenting ability in lactic streptococci . Appl. Environ. Microbiol. 54 : 1951 - 1956 .
Norgren , M. , and J. R. Scott . 1991 . The presence of conjugative transposon Tn 916 in the recipient strain does not impede transfer of a second copy of the element . J. Bacteriol. 173 : 319 - 324 .
Rauch , P. J. G. , M. M. Beerthuyzen , and W. M. de Vos . 1990 . Nucleotide sequence of IS904 from Lactococcus lactis subsp. lactis strain NIZO R5 . Nucleic Acids Res. 18 : 4253 - 4254 .
Rauch P. J. G. M. M. Beerthuyzen and W. M. de Vos. 1991. Molecular analysis and evolution of conjugative transposons encoding nisin production and sucrose fermentation in Lactococcus lactis p. 243-249. In G. Jung and H.-G. Sahl (ed.) Nisin and novel lantibiotics. Escom Leiden The Netherlands.
Rauch P. J. G. and W. M. de Vos. 1990. Molecular analysis of the Lactococcus lactis nisin-sucrose conjugative transposon abstr. A/46 p. 23. Program Abstr. 3rd Int. Am. Soc. Microbiol. Conference on Streptococcal Genetics. American Society for Microbiology Washington D.C.
Rauch P. J. G. and W. M. de Vos. Submitted for publication.
Rauch P. J. G. and W. M. de Vos. Unpublished data.
Reed , K. C. , and D. A. Mann . 1985 . Rapid transfer of DNA from agarose gels to nylon membranes . Nucleic Acids Res. 13 : 7207 - 7221 .
Sambrook J. E. F. Fritsch and T. Maniatis. 1989. Molecular cloning: a laboratory manual 2nd ed. Cold Spring Harbor Laboratory Cold Spring Harbor N.Y.
Sanger , F. , S. Nicklen , and A. R. Coulson . 1977 . DNA sequencing with chain-terminating inhibitors . Proc. Natl. Acad. Sci. USA 74 : 5463 - 5467 .
Steele , J. L. , and L. L. McKay . 1986 . Partial characterization of the genetic basis for sucrose metabolism and nisin production in Streptococcus lactis . Appl. Environ. Microbiol. 51 : 57 - 64 .
Steele , J. L. , and L. L. McKay . 1989 . Conjugal transfer of genetic material by Lactococcus lactis subsp. lactis 11007 . Plasmid 22 : 32 - 43 .
Steen , M. T. , Y. J. Chung , and J. N. Hansen . 1991 . Characterization of the nisin gene as part of a polycistronic operon in the chromosome of Lactococcus lactis ATCC 11454 . Appl. Environ. Microbiol. 57 : 1181 - 1188 .
Thompson , J. , R. J. Harr , J. A. Donkersloot . 1990 . N5-(L-1- carboxyethyl)-L-ornithine: distribution, constitutivity, and regulation . Curr. Microbiol. 20 : 239 - 244 .
Traimer , J. , and G. G. Fowler . 1964 . Estimation of nisin in foods . J. Sci. Food Agric. 15 : 522 - 528 .
Trien-Cuot P. C. Poyart-Salmeron C. Carlier and P. Courvalin. 1991. Molecular dissection of the transposition mechanism of conjugative transposons from gram-positive cocci p. 21-27. In G. M. Dunny P. P. Cleary and L. L. McKay (ed.) Genetics and molecular biology of streptococci lactococci and enterococci. American Society for Microbiology Washington D.C.
Van Alen-Boerrigter , I. J. , R. Baankreis , and W. M. de Vos . 1991 . Characterization and overexpression of the Lactococcus lactispepN gene and localization of its product, aminopeptidase N . Appl. Environ. Microbiol. 57 : 2555 - 2561 .
Vjayakumar M. N. P. Ayoubi and A. 0. Kilic. 1991. Organization of Tn 5253 the pneumococcal Qt(cat tet) BM6001 element p. 49-53. In G. M. Dunny P. P. Cleary and L. L. McKay (ed.) Genetics and molecular biology of streptococci lactococci and enterococci. American Society for Microbiology Washington D.C.
Viayakumar , M. N. , S. D. Priebe , and W. R. Guild . 1986 . Structure of a conjugative element in Streptococcus pneumoniae . J. Bacteriol. 166 : 978 - 984 .
Vos , P. , M. van Asseldonk , F. van Jeveren , R. Siezen , G. Simons , and W. M. de Vos . 1989 . A maturation protein is essential for production of active forms of Lactococcus lactis SK11 serine proteinase located in or secreted from the cell envelope . J. Bacteriol. 171 : 2795 - 2802 .
Woolley , R. C. , A. Pennock , R. J. Ashton , A. Davies , and M. Young . 1989 . Transfer of TnJS45 and Tn 916 to Clostridium acetobutylicum . Plasmid 22 : 169 - 174 .