Characterization of a "mobilization cassette" in transposon Tn4399 from Bacteroides fragilis.
Derivatives of nonconjugal plasmids that carry Tn4399, a transposon isolated from Bacteroides fragilis, can be mobilized for transfer by the broad-host-range IncP plasmids pRK231 or R751 in Escherichia coli. To characterize regions of Tn4399 involved in mobilization, we have isolated and analyzed subcloned fragments of Tn4399 in E. coli, as well as mutations within the element. We have identified a "mobilization cassette" within a 2.8-kb region of Tn4399 which, when cloned into mobilization-deficient plasmids, allows these plasmids to be mobilized in trans by the IncP plasmids pRK231 and R751. The 2.8-kb region has been sequenced, and several open reading frames have been identified. Mutants defective in two genes, designated mocA and mocB, coding for deduced products of 36.4 and 16.4 kDa, respectively, cannot be mobilized by either IncP plasmid; these mutants can be complemented in the presence of the respective wild-type genes in trans. This suggests that the putative MocA and MocB proteins have a role in the mobilization process. The 36.4-kDa MocA protein contains a 14-amino-acid sequence which is closely related to a highly conserved motif within DNA relaxases encoded by a wide variety of conjugal or mobilizable plasmids. Subcloning experiments also lead to the localization of an oriT region within a 199-bp fragment, internal to the mobilization cassette.
Murphy
C G
CG
Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts 02111.
Malamy
M H
MH
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001441:Bacteroides fragilis / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D003227:Conjugation, Genetic / Q000235:genetics*; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010957:Plasmids; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; C083182:MocA protein, Bacteroides fragilis; C083184:MocB protein, Bacteroides fragilis