Please refer to https://www.mikrobio.uni-kiel.de/de/ag-bramkamp and contact Marc <email@example.com> for further information on the project.
Evolution of DNA-segregation machineries in prokaryotes
A hallmark of life is the replication and inheritance of genetic material. Prokaryotic cells, bacteria and archaea, harbor often chromosomal and extra-chromosomal replicons that coordinate their replication and segregation with the cell cycle. To ensure faithful segregation, active transport systems have evolved that specifically bind to individual replicons and mediate segregation. These systems usually consist of a centromere-binding protein that recognizes cis-acting DNA sequences (the centromeric region) and an ATP-hydrolyzing motor protein. The classical system is the ParBAS system that has been identified on many bacterial chromosomes and plasmids. ParB is a CTP-binding molecular switch that binds to parS sites and ParA is a Walker-type ATPase that uses the existing chromosome as a track to segregate either plasmids or the chromosomal origin of replication. Despite detailed knowledge about bacterial chromosome replication and segregation, these processes in Archaea remain obscure. Haloarchaea, a class of the Euryarchaeota, harbor many plasmids and are usually polyploid. We have identified plasmid specific segregation systems, with distant homology to the described ParABS system in bacteria. However, the DNA-binding proteins are evolutionary unrelated to the bacterial ParB proteins and the ParA proteins fall into at least two lineages. Therefore, we designated these system haloarchaeal partition A and B, HpaA/ HpaB.
Students will use fluorescence microscopy, to study the subcellular localization of HpaAB. Dynamics of the HpaAB proteins will be analyzed by single-molecule localization microscopy. Therefore, Applicants will receive an in-depth training in super-resolution microscopy and subsequent data analysis. Furthermore, DNA-binding sites of HpaB will be identified by CHIP-Seq analysis and electrophoretic mobility shift assays. Additionally, the HpaAB system will be analyzed biochemically. Homologous expression and purification of HpaAB will be performed and proteins will be biochemically analyzed. Finally, the dynamics of the system will be fully reconstructed in vitro. This broad range of techniques gives a unique opportunity to get professional insight into modern biochemistry, cell biology and genetics.
- Böhm K, Giacomelli G, Schmidt A, Imhof A, Koszul R, Marbouty M, Bramkamp M. (2020) Chromosome organization by a conserved condensin-ParB system in the actinobacterium Corynebacterium glutamiucm. Nature Comm., 11, 1485
- Böhm K, Meyer F, Rhomberg A, Kalinowski J, Donovan C, Bramkamp M. (2017) Novel Chromosome Organization Pattern in Actinomycetales-Overlapping Replication Cycles Combined with Diploidy mBio. 8(3):e00511-17. doi: 10.1128/mBio.00511-17.