*Author for correspondence. Correspondence should be addressed to: Department of Molecular Genetics, Institute of Chemical Physics and Biophysics, Akadeemia tee 23, EE0026 Tallinn, Republic of Estonia. Tel. +372 2529247; fax +372 2529579.
† Present address: Department of Biochemistry, Tallinn Technical University, Ehitajate tee 5, EH0108 Tallinn, Republic of Estonia.
SUMMARY: Two new segregants, PPW1-1 and PPW161-1, of Pseudomonas putida were isolated from the stock cultures PaW85(pWW0) and PaW85(pWW0-161). Strain PPW1-1 had lost its ability to grow on m-xylene but was able to grow on m-toluate. A deletion of the left-hand of transposon Tn4651, including the upper-operon genes, had taken place in plasmid pWWOmut11, isolated from strain PPW1-1. Additional deletions were observed in pWWOmut1 after ‘benzoate-curing’ (plasmids pWW0mut15, pWWOmut19, pWW0mut27). The genes of the upper-operon and beginning of the meta-operon were deleted from pWW0-161mut1, isolated from strain PPW161-1. Despite this deletion, cells of PPW161-1 grew on all normal TOL plasmid substrates. The Tol+ phenotype was stable in cells of PPW161-1 growing on benzoate. We propose that this is because in cells of strain PPW161-1 the catabolic genes deleted from pWW0-161mut1 were integrated into the chromosome at the site where the (chromosomally encoded) ortho-pathway genes are located, resulting in the inability of the cells to use this pathway.
BayleyS.A.,
DUGGLEBYC.J.,
WORSEYM.J.,
WILLIAMSP.A.,
HARDYK.G.,
BRODAP.1977; Two modes of loss of the Tol function from Pseudomonas putida mt-2.. Molecular and General Genetics 154:203–204
BradfordM.M.1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.. Analytical Biochemistry 72:248–254
DowningR.G.,
DUGGLEBYC.J.,
VillemsR.,
BrodaP.1979; An endonuclease cleavage map of the plasmid pWW0-8, a derivative of the TOL plasmid of Pseudomonas putida mt-2.. Molecular and General Genetics 168:97–99
FeistC.F.,
HegemanG.D.1969; Phenol and benzoate metabolism by Pseudomonas putida: regulation of tangential path-way.. Journal of Bacteriology 100:869–877
FranklinF.C.H.,
BAGDASARIANM.,
BAGDASARIANM.M.,
TIMMISK.N.1981; Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta-cleavage pathway.. Proceedings of the National Academy of Sciences of the United States of America787458–7462
HarayamaS.,
RekikR.,
WubblotsM.,
RoseK.,
LeppikR.A.,
TimmisK.N.1989; Characterization of five genes in the upper-pathway operon of TOL plasmid pWW0 from Pseudomonas putida and identification of the gene products.. Journal of Bacteriology 171:5048–5055
KunzD.A.,
ChapmanP.J.1981; Isolation and characterization of spontaneously occurring TOL plasmid mutants of Pseudomonas putida HS1.. Journal of Bacteriology 146:952–964
LebensM.R.,
WilliamsP.A.1985; Complementation of deletion and insertion mutants of TOL plasmid pWWO: regulatory implications and location of xylC gene.. Journal of General Microbiology 131:3261–3269
LehrbachP.R.,
WardJ.,
MeulienP.,
BrodaP.1982; Physical mapping of TOL plasmids pWW0 and pND2 and various R plasmid-TOL derivatives from Pseudomonas sp.. Journal of Bacteriology 152:1280–1283
MÄEA.A.,
HabichtYA.K.,
NURKA.E.,
HeinaruA.L.1991; Transposons Tn4652 and Tn3614 of the TOL plasmid pWW0 are involved in conjugal mobilization of chromosomally located catabolic cam-operons.. Genetika 27:773–782
MeulienP.,
BrodaP.1982; Identification of chromosomally integrated TOL DNA in cured derivatives of Pseudomonas putida PaWl.. Journal of Bacteriology 152:1280–1283
MeulienA.P.,
DowningR.G.,
BrodaP.1981; Excision of the 40 kb segment of the TOL plasmid from Pseudomonas putida mt-2 involves direct repeats.. Molecular and General Genetics 184:97–101
RamosJ.L.,
MermodN.,
TimmisK.N.1987; Regulatory circuits controlling transcription of TOL plasmid operon encoding metacleavage pathway for degradation of alkylbenzoates by Pseudomonas.. Molecular Microbiology 1:293–300
ShawL.E.,
WilliamsP.A.1988; Physical and functional mapping of two cointegrate plasmids derived from RP4 and TOL plasmid pDKl.. Journal of General Microbiology 134:2463–2474
SinclairM.I.,
MaxwellP.C.,
LyonB.R.,
HollowayB.W.1986; Chromosomal location of TOL plasmid DNA in Pseudomonas putida.. Journal of Bacteriology 168:1302–1308
TsudoM.,
IinoT.1988; Identification and characterization of Tn4653, a transposon covering the toluene transposon Tn4651 on TOL plasmid pWW0.. Molecular and General Genetics 213:72–77
WilliamsP.A.,
MurrayK.1974; Metabolism of benzoate and the methylbenzoates by Pseudomonas putida (arvilla) mt-2: evidence for the existence of a TOL plasmid.. Journal of Bacteriology 120:416–423
WilliamsP.A.,
TaylorS.D.,
GIBBL.E.1988; Loss of the toluene-xylene catabolic genes of TOL plasmid pWW0 during growth of Pseudomonas putida on benzoate is due to a selective growth advantage of ‘cured’ segregants.. Journal of General Microbiology 134:2039–2048
WorseyM.J.,
WilliamsP.A.1975; Metabolism of toluene and xylenes by Pseudomonas putida (arvilla) mt-2; evidence for a new function of the TOL plasmid.. Journal of Bacteriology 127:7–13