Dynamic assembly, disassembly and bundling of the bacterial cell division protein FtsZ and YgfE (ZapA)

• Main Author: Cheng, Xi
• Published: University of Warwick 2011
• Subjects: 540; QR Microbiology
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.560236

The protein FtsZ is a tubulin-like GTPase, which plays an essential role in prokaryotic cell division. In vivo it assembles into a dynamic ring (the Z-ring) at the future site of cell division on the inner surface of the cytoplasmatic membrane. The Z-ring then serves as a scaffold which recruits all other division proteins to form the cytokinetic machinery. The constriction of the ring facilitates the separation of two daughter cells. In vitro, FtsZ polymerizes in the presence of GTP to form single-stranded protofilaments. It is assumed that FtsZ association and assembly reactions studied in vitro will play an important role in understanding the mechanism of Z-ring assembly and disassembly in vivo. In this work, we therefore have studied the dynamics of FtsZ polymerization in vitro, especially the bundling induced with YgfE. YgfE, the putative Escherichia coli ZapA orthologue, is one of these division proteins recruited by FtsZ. It acts to enhance lateral associations between FtsZ fibres to form larger bundles. In this study, we have demonstrated that YgfE exists as a tetramer in solution at the concentrations reported in this study, and the bundling activity is exerted more efficiently on preformed FtsZ protofilaments. We also investigate the importance of the tetramerisation of YgfE on function. A number of mutant forms of ZapA were generated with the aim of disrupting the dimer:dimer interface. Using those mutants we show that tetramerisation is a requirement for both FtsZ bundling, and GTPase modulation activities. In addition, a novel technique, continuous channel flow linear dichroism (LD) spectroscopy, has been first developed in this work. LD is a simple spectroscopy technique for structural characterization of long biomacromolecules in solutions. Continuous channel flow Flow LD has overcome the limitation of Couette flow LD due to the time required to assemble and fill the cell.