Item – Theses Canada

​
OCLC number
1007021981
Link(s) to full text
LAC copy
LAC copy
Author
Surtees, Jennifer A.(Jennifer Anne),1971-
Title
P1 ParB : the structural and functional domains of the P1 plasmid centromere-binding protein.
Degree
Ph. D. -- University of Toronto, 2001
Publisher
Ottawa : National Library of Canada = Bibliothèque nationale du Canada, [2002]
Description
2 microfiches
Notes
Includes bibliographical references.
Abstract
The P1 prophage plasmid is stably maintained as a unit-copy plasmid in ' Escherichia coli'. Faithful maintenance of P1 requires an active partition system, 'par', that is encoded by the plasmid. The ' par' operon consists of two transacting factors, 'parA' and 'parB', and a centromere-like site called 'parS '. ParB is a multifunctional protein. It interacts with and stimulates the biochemical activities of ParA. ParB is a dimer in solution. Finally, ParB binds specifically to 'parS', along with the ' E. coli' integration host factor (IHF) to form a high affinity nucleoprotein complex called the partition complex, which mediates partition. ParB and IHF stimulate each other's DNA binding activity. In this thesis I examine the structural and functional domains of ParB. Using protein fragments, I show that ParB contains two independent multimerization domains, one at its C-terminus and one at its N-terminus. I also show that the extreme N-terminus of ParB is required for ParA-ParB interactions. By limited proteolytic digestion, I show that ParB has a distinct domain structure, with the C-terminal dimerization domain at its core. I have examined the architecture of the partition complex by investigating the DNA binding activity of various ParB fragments. A single dimer of ParB is sufficient for partition complex formation. The first 141 residues of ParB are dispensable for the formation of the partition complex. A fragment missing only the last 16 amino acids of ParB binds specifically to 'parS', but binding is weak and no longer stimulated by IHF. The ability of IHF to stimulate ParB binding to 'parS' correlates with C-terminal dimerization. Using full and partial 'parS ' sites, I have found that two regions of ParB, one in the centre and the other near the C-terminus, interact with distinct sequences within ' parS'. I propose a model of how the ParB dimer binds 'parS ' to form the minimal partition complex.
ISBN
0612590267
9780612590267