Characterizing ParA2, an essential protein for Vibrio cholerae chromosome segregation.
Hui, Monica.
2011
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Abstract: Partitioning, or par, loci were initially identified on low copy plasmids as elements essential for plasmid segregation and inheritance. However, it is now apparent that many bacterial chromosomes harbor par loci as well. The role of chromosomal par loci remains incompletely understood, but their homology to plasmid par loci has led to speculation that chromosomal par loci may play a key... read morerole in mediating chromosome segregation. The genome of Vibrio cholerae, the causative agent of cholera, is divided between two chromosomes, each of which encodes its own par locus. Prior studies have shown that the parAB2 locus is essential for the proper segregation of chromosome II. In order to begin to elucidate the molecular mechanisms by which parAB2 mediates V. cholerae chromosome II segregation, I focused on ParA2, the protein thought to drive the partitioning process. In vitro, I found that ParA2 shares biochemical characteristics with other members of the ParA family of partitioning proteins. However, unlike most ParA family proteins, which polymerize in the presence of ATP, ParA2 only formed higher ordered filamentous structures on DNA. Three-dimensional reconstructions of these filaments revealed that ParA2 binds DNA to form unusual left-handed nucleoprotein filaments that are bipolar and symmetric. These filaments do not distort the DNA helix bound in the interior. Subsequent in vivo studies using ParA2 fluorescent protein fusions showed that this partitioning protein is dynamic and nucleoid associated. Strikingly, ParA2 is often found between a ParB2-parS2 centromere complex and the cell pole towards which that complex is moving. Together, my findings suggest that ParA2 forms bipolar filaments over the nucleoid and utilizes a pulling mechanism to mediate partitioning of chromosome II.
Thesis (Ph.D.)--Tufts University, 2011.
Submitted to the Dept. of Molecular Microbiology.
Advisor: Matthew Waldor.
Committee: Ekaterina Heldwein, Andrew Wright, Linc Sonenshein, Ralph Isberg, and David Rudner.
Keyword: Microbiology.read less - ID:
- bc386w71w
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