M Adamczyk1, E Lewicka2, R Szatkowska3, H Nieznanska4, J Ludwiczak5,6, M Jasiński5, S Dunin-Horkawicz5, E Sitkiewicz7, B Swiderska7, G Goch2, G Jagura-Burdzy2. 1. Warsaw University of Technology, Faculty of Chemistry, Chair of Drug and Cosmetics Biotechnology, Noakowskiego 3, 00-664, Warsaw, Poland. madamczyk@ch.pw.edu.pl. 2. Department of Microbial Biochemistry, Institute of Biochemistry and Biophysics PAS, Pawinskiego 5a, 02-106, Warsaw, Poland. 3. Warsaw University of Technology, Faculty of Chemistry, Chair of Drug and Cosmetics Biotechnology, Noakowskiego 3, 00-664, Warsaw, Poland. 4. Nencki Institute of Experimental Biology PAS, Laboratory of Electron Microscopy, Pasteura 3, 02-093, Warsaw, Poland. 5. University of Warsaw, Centre of New Technologies, Laboratory of Structural Bioinformatics, 02-097, Warsaw, Poland. 6. Nencki Institute of Experimental Biology, Laboratory of Bioinformatics, Pasteura 3, 02-093, Warsaw, Poland. 7. Mass Spectrometry Laboratory, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawinskiego 5a, 02-106, Warsaw, Poland.
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.
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.