The Min system ensures that gram-negative bacteria undergo symmetric cell division. The three Min proteins, MinC, MinD, and MinE, display a dynamic pattern of subcellular organization on the inner cell membrane that directs division proteins to the mid-cell. This process is driven by the ATPase activity of MinD that is stimulated through its interaction s with Min E. A recent structure of MinE in complex with MinD suggests that MinE undergoes a dramatic conformational change to allow MinD - binding residues to be released from the MinE hydrophobic core. However, this structure used a MinE mutant designed to favor this conformational change, raising questions regarding the mechanism by which wild - type MinE can undergo this transition in vivo. One potential scenario that might explain this structural change involves a recently discovered interaction between MinE and the membrane surface. To investigate the possibility that lipid binding could induce this structural transition in MinE, circular dichroism and enzyme kinetics studies were carried out. These studies were also done on MinE mutants designed to either eliminate membrane binding or induce the conformational change involved in MinD - binding. The results demonstrated that a membrane induced conformational change does occur, and requires the presence of a key lipid - binding region at the N - terminus. However, removal of this sequence failed to alter the kinetics of MinE - stimulated MinD - catalyzed ATP hydrolysis. Overall, our results provide a step forward in our understanding of the role of the interaction between MinE and the membrane in the Min system, but also highlight the need for additional investigation before this system might be used as a novel antibiotic target for pathogenic, gram - negative bacteria such as Neisseria gonorrhoeae.

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Implementation and analysis of mathematical models can serve as a powerful tool in understanding how intracellular processes in bacteria affect the bacterial phenotype. In this thesis I have implemented and analysed models of a number of different parts of the bacterium E. coli in order to understand these types of connections. I have also developed new tools for analysis of stochastic reaction-diffusion models. Resistance mutations in the E. coli ribosomes make the bacteria less susceptible to treatment with the antibiotic drug erythromycin compared to bacteria carrying wildtype ribosomes. The effect is dependent on efficient drug efflux pumps. In the absence of pumps for erythromycin, there is no difference in growth between wildtype and drug target resistant bacteria. I present a model explaining this unexpected phenotype, and also give the conditions for its occurrence. Stochastic fluctuations in gene expression in bacteria, such as E. coli, result in stochastic fluctuations in biosynthesis pathways. I have characterised the effect of stochastic fluctuations in the parallel biosynthesis pathways of amino acids. I show how the average protein synthesis rate decreases with an increasing number of fluctuating amino acid production pathways. I further show how the cell can remedy this problem by using sensitive feedback control of transcription, and by optimising its expression levels of amino acid biosynthetic enzymes. The pole-to-pole oscillations of the Min-proteins in E. coli are required for accurate mid-cell division. The phenotype of the Min-oscillations is altered in three different mutants: filamentous cells, round cells and cells with changed membrane lipid composition. I have shown that the wildtype and mutant phenotypes can be explained using a stochastic reaction-diffusion model. In E. coli, the transcription elongation rate on the ribosmal RNA operon increases with increasing transcription initiation rate. In addition, the polymerase density varies along the ribosomal RNA operons. I present a DNA sequence dependent model that explains the transcription elongation rate speed-up, and also the density variation along the ribosomal operons. Both phenomena are explained by the RNA polymerase backtracking on the DNA. / Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 715