Molecular mechanism of the dynamic association of MinD with the membrane in dividing bacteria

Date of Completion

January 2004

Keywords

Biology, Microbiology|Chemistry, Biochemistry

Degree

Ph.D.

Abstract

Cytokinesis in bacteria is a complex process subject to precise spatiotemporal regulations. In E. coli, genetically and morphologically equivalent progeny are produced by binary fission at midcell. Formation of the septum is dependent on the essential protein, FtsZ, which assembles into a ring structure (the Z-ring) at the future plane of division. Asymmetric divisions, which produce chromosomeless minicells and short filaments, are avoided through the coordinated actions of the Min proteins (MinC, MinD, and MinE) which exhibit a rapid pole-to-pole oscillation. This oscillation serves to discourage polar divisions by disrupting FtsZ ring formation at sites other than at midcell. Failure to oscillate, or alterations to the frequency of oscillation, lead to deregulation of Z-ring placement and the formation of minicells. The oscillation is biphasic, consisting of an extended polar membrane occupancy period and a shorter, apparently cytoplasmic migration period. ^ In this doctoral thesis, we investigate the molecular mechanism controlling the switch between the membrane bound and cytoplasmic phases of Min oscillation. Membrane association of the Min proteins is orchestrated by the amphitropic MinD protein. We find that a short (8–12 residue) C-terminal motif is necessary and sufficient for MinD membrane binding. This motif, which we have termed the membrane targeting sequence (MTS), is conserved in MinD homologues across eubacteria, archaea, and plastids. The MTS has a propensity to form an amphipathic α-helix. Mutations within the MTS that disrupt either the helicity or its amphipathic nature result in MinD being incapable of localizing properly to the inner membrane, rendering the entire Min system non-functional. We determine that the presence of anionic phospholipids in the membrane is essential for the MTS to bind. Furthermore, we provide evidence that the MinD MTSs from different organisms have evolved to recognize specific phospholipids within that organism. We suggest that this specificity, in addition to ATP hydrolysis, regulate MinD function. Intriguingly, the MTS alone is capable of targeting MinC to the membrane to cause a division block. Finally, based on our results, we propose a new model that explains how the MTS is intimately involved in MinD membrane attachment and polymer growth. ^

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