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Abstract Background DNA binding KfrA-type proteins of broad-host-range bacterial plasmids belonging to IncP-1 and IncU incompatibility groups are characterized by globular N-terminal head domains and long alpha-helical coiled-coil tails. They have been shown to act as transcriptional auto-regulators. Results This study was focused on two members of the growing family of KfrA-type proteins encoded by the broad-host-range plasmids, R751 of IncP-1β and RA3 of IncU groups. Comparative in vitro and in silico studies on KfrAR751 and KfrARA3 confirmed their similar biophysical properties despite low conservation of the amino acid sequences. They form a wide range of oligomeric forms in vitro and, in the presence of their cognate DNA binding sites, they polymerize into the higher order filaments visualized as “threads” by negative staining electron microscopy. The studies revealed also temperature-dependent changes in the coiled-coil segment of KfrA proteins that is involved in the stabilization of dimers required for DNA interactions. Conclusion KfrAR751 and KfrARA3 are structural homologues. We postulate that KfrA type proteins have moonlighting activity. They not only act as transcriptional auto-regulators but form cytoskeletal structures, which might facilitate plasmid DNA delivery and positioning in the cells before cell division, involving thermal energy.

PromA plasmids are broad host range (BHR) plasmids, which are often cryptic and hence have an uncertain ecological role. We present three novel PromA γ plasmids which carry genes associated with degradation of the phenylurea herbicide linuron, two of which originated from unrelated Hydrogenophaga hosts isolated from different environments (pPBL-H3-2 and pBPS33-2), and one (pEN1) which was exogenously captured from an on-farm biopurification system (BPS). Hydrogenophaga sp. plasmid pBPS33-2 carries all three necessary gene clusters (hylA, dca, ccd) determining the three main steps for conversion of linuron to Krebs cycle intermediates, while pEN1 only determines the initial linuron hydrolysis step. Hydrogenophaga sp. plasmid pPBL-H3-2 exists as two variants, both containing ccd but with the hylA and dca gene modules interchanged between each other at exactly the same location. Linuron catabolic gene clusters that determine the same step were identical on all plasmids, encompassed in differently arranged constellations and characterized by the presence of multiple IS1071 elements. In all plasmids except pEN1, the insertion spot of the catabolic genes in the PromA γ plasmids was the same. Highly similar PromA plasmids carrying the linuron degrading gene cargo at the same insertion spot were previously identified in linuron degrading Variovorax sp. Interestingly, in both Hydrogenophaga populations not every PromA plasmid copy carries catabolic genes. The results indicate that PromA plasmids are important vehicles of linuron catabolic gene dissemination, rather than being cryptic and only important for the mobilization of other plasmids.

Background: DNA binding KfrA-type proteins of broad-host-range bacterial plasmids belonging to IncP-1 and IncU incompatibility groups are characterized by globular N-terminal head domains and long alpha-helical coiled-coil tails. They have been shown to act as transcriptional auto-regulators. Results: This study was focused on two members of the growing family of KfrA-type proteins encoded by the broad-host-range plasmids, R751 of IncP-1β and RA3 of IncU groups. Comparative in vitro and in silico studies on KfrAR751 and KfrARA3 confirmed their similar biophysical properties despite low conservation of the amino acid sequences. They form a wide range of oligomeric forms in vitro and, in the presence of their cognate DNA binding sites, they polymerize into the higher order filaments visualized as "threads" by negative staining electron microscopy. The studies revealed also temperature-dependent changes in the coiled-coil segment of KfrA proteins that is involved in the stabilization of dimers required for DNA interactions. Conclusion: KfrAR751 and KfrARA3 are structural homologues. We postulate that KfrA type proteins have moonlighting activity. They not only act as transcriptional auto-regulators but form cytoskeletal structures, which might facilitate plasmid DNA delivery and positioning in the cells before cell division, involving thermal energy. [ABSTRACT FROM AUTHOR]

PromA plasmids are broad host range (BHR) plasmids, which are often cryptic and hence have an uncertain ecological role. We present three novel PromA γ plasmids which carry genes associated with degradation of the phenylurea herbicide linuron, two of which originated from unrelated Hydrogenophaga hosts isolated from different environments (pPBL-H3-2 and pBPS33-2), and one (pEN1) which was exogenously captured from an on-farm biopurification system (BPS). Hydrogenophaga sp. plasmid pBPS33-2 carries all three necessary gene clusters (hylA, dca, ccd) determining the three main steps for conversion of linuron to Krebs cycle intermediates, while pEN1 only determines the initial linuron hydrolysis step. Hydrogenophaga sp. plasmid pPBL-H3-2 exists as two variants, both containing ccd but with the hylA and dca gene modules interchanged between each other at exactly the same location. Linuron catabolic gene clusters that determine the same step were identical on all plasmids, encompassed in differently arranged constellations and characterized by the presence of multiple IS 1071 elements. In all plasmids except pEN1, the insertion spot of the catabolic genes in the PromA γ plasmids was the same. Highly similar PromA plasmids carrying the linuron degrading gene cargo at the same insertion spot were previously identified in linuron degrading Variovorax sp. Interestingly, in both Hydrogenophaga populations not every PromA plasmid copy carries catabolic genes. The results indicate that PromA plasmids are important vehicles of linuron catabolic gene dissemination, rather than being cryptic and only important for the mobilization of other plasmids. [ABSTRACT FROM AUTHOR]