Identification and Characterization of Tn 4656 , a Novel Class II Transposon Carrying a Set of Toluene-Degrading Genes from TOL Plasmid pWW53

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Abstract
It has been reported that the toluene-degrading ( xyl ) genes from Pseudomonas putida plasmid pWW53 are able to translocate to broad-host-range drug resistance plasmid RP4, and pWW53-4 is one of the smallest RP4 derivatives (H. Keil, S. Keil, R. W. Pickup, and P. A. Williams, J. Bacteriol. 164:887–895, 1985). Our investigation of pWW53-4 in this study demonstrated that such a translocated region that is 39 kb long is a transposon. This mobile element, Tn 4656 , was classified as a class II transposon since its transposition occurred by a two-step process: transposase (TnpA)-mediated formation of the cointegrate and resolvase (TnpR)-mediated site-specific resolution of the cointegrate at the two copies of the res site. The Tn 4656 TnpA and TnpR functions encoded in the rightmost 4-kb region were found to be exchangeable with those specified by other Tn 1721 -related class II transposons, including another toluene transposon, Tn 4653 . Sequence analysis of the transposition-related genes and sites of Tn 4656 also supported the hypothesis that this transposon is closely related to the Tn 1721 -related transposons. The lower transposition frequency of Tn 4656 has been suggested to be due to the unique nucleotide sequence of one of the terminal 39-bp inverted repeats.