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Abstract
Bacillus subtilis SpoIIIE is a member of the FtsK/SpoIIIE family of double-stranded DNA (dsDNA) transporters that are involved in a wide variety of processes in many species, including: conjugation, DNA packaging of eukaryotic and bacterial viruses, chromosome segregation in bacteria, and DNA maintenance in archae. SpoIIIE moves a trapped chromosome across an asymmetric division septum into the forespore during sporulation. To further understand how members of the FtsK/SpoIIIE family interact with their substrates, we investigated the role of the &agr; domain of SpoIIIE during sporulation in B. subtilis, and investigated how SpoIIIE interacts with its substrate DNA during transport by taking advantage of the non-translocating mutant: SpoIIIE36.
The work described in this thesis tells a story that begins with the characterization of a mutation of SpoIIIE (SpoIIIE36) and ends by beginning to map the interactions between SpoIIIE and its substrate DNA. Here we find that the mutation V429M disrupts the ability of SpoIIIE to form an oligomer in vivo and in vitro. Additionally, we find that the &agr; domain of SpoIIIE plays a role in maintenance of the oligomeric state. Finally, we find that various positions within the pore loops of SpoIIIE are capable of interacting with the substrate DNA. These results begin to illuminate a mechanism for how the family of FtsK/SpoIIIE DNA transporters interact with their substrates during translocation.
