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1 The Institute of Biochemistry and Biophysics, Polish Academy of Sciences (PAS), 02-106 Warsaw, Pawinskiego 5A, Poland
2 Central Institute of Labour Protection, National Research Institute, 00-701 Warsaw, Czerniakowska 16, Poland
3 School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK
Correspondence
Grazyna Jagura-Burdzy
gjburdzy{at}poczta.ibb.waw.pl
The kfrA gene of the IncP-1 broad-host-range plasmids is the best-studied member of a growing gene family that shows strong linkage to the minimal replicon of many low-copy-number plasmids. KfrA is a DNA binding protein with a long, alpha-helical, coiled-coil tail. Studying IncP-1
plasmid R751, evidence is presented that kfrA and its downstream genes upf54.8 and upf54.4 were organized in a tricistronic operon (renamed here kfrA kfrB kfrC), expressed from autoregulated kfrAp, that was also repressed by KorA and KorB. KfrA, KfrB and KfrC interacted and may have formed a multi-protein complex. Inactivation of either kfrA or kfrB in R751 resulted in long-term accumulation of plasmid-negative bacteria, whereas wild-type R751 itself persisted without selection. Immunofluorescence studies showed that KfrAR751 formed plasmid-associated foci, and deletion of the C terminus of KfrA caused plasmid R751
C2kfrA foci to disperse and mislocalize. Thus, the KfrABC complex may be an important component in the organization and control of the plasmid clusters that seem to form the segregating unit in bacterial cells. The studied operon is therefore part of the set of functions needed for R751 to function as an efficient vehicle for maintenance and spread of genes in Gram-negative bacteria.
The DNA sequence and predicted amino acid sequence of the R751 region containing kfrA–upf54.8 and the 5' end of upf54.4 are available as supplementary data with the online version of this paper.
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