Ether-lipid membrane engineering of Escherichia coli

The thesis describes the reconstitution of the complete ether lipid biosynthetic pathway both in vitro and in vivo. The study identified a novel enzyme in the archaeal lipids biosynthesis, namely the CDP-archaeol synthase and revealed a high promiscuity of bacterial enzymes for the attachment of the polar head groups. The work also sheds some light on the significance of some evolutionary theories. The obtainment of a bacterial strain with a heterochiral mixed membrane showed the coexistence of the two different lipid species contrasting the previous hypothesis of chemical instability of such membrane which pushed the evolution of bacterial and archaeal organisms towards a more stable homochiral membranes. Thus, the coexistence of bacterial and archaeal lipids in a living cell suggested the presence of a common ancestor with a mixed membrane from which organisms evolved. Further, the different substrate specificity found in the archaeal GGGPS and in the bacterial PlsB raises new possibilities about the initial insurgence of archaeal organisms, followed by the differentiation into bacteria. In this way the trigger that caused the segregation from the primordial cell can be seen as the appearance of stereoselective enzymes such as PlsB which pushed the differentiation of bacterial organisms from the ancient cell. The latter one then further evolved towards an archaeal organism with the conserved capability to live in extreme environments but with an evolved and specific membrane lipid composition. Further, the work has potential biotechnological implications as the re-programming of the lipid composition of E. coli resulted in some increase in robustness. Thus, when further tuned, this may be seen as a general methodology for other microorganisms of industrial relevance when aspects of solvent tolerance are addressed. It would not be possible to inflict such radical alteration of the lipidome by mutagenesis and selection