Fatty Acid Insertion Mechanism During Membrane Synthesis In Escherichia Coli

Cell membrane is an essential part of a cell,which bases on phospholipid bilayer.And the main components of membranes are lipid and protein.The studies of cell membrane synthesis in Escherichia coli mostly focused on the physiological function of membrane synthases,and there was not any report on the mechanism of fatty acid insertion during the process of membrane synthesis or the localization of membrane synthesis.In order to explore these issues in Escherichia coli,we conducted a series of researches.As 80%of the membrane phospholipids in E.coli are phosphatidylethanolamine(PE),we focus on the synthesis of PE,fusing the 10 key enzymes involved in PE synthesis with green fluorescent protein(EGFP)or red fluorescent protein(mCherry)and expressing the these fusion proteins in E.coli.We used laser confocal fluorescence microscopy to obtain localization information of these fusion proteins.When expressed at high level,EGFP-FabA,EGFP-FabB,EGFP-FabI,EGFP-FabG,EGFP-PlsB and EGFP-PssA accumulated in poles and septum.While EGFP-FabD,EGFP-FabF,EGFP-CdsA and EGFP-PSD are uniformly dispersed in the cytoplasm or cell membrane at varies expression levels.Time-lapse images showed that EGFP-PlsB,a key protein in the synthesis pathway of PE,moved along with the growth and division of the cell and accumulated in the septum before cell division.We speculate that fatty acids inserted in septum and the PE is mostly synthesized in the septum and poles,and are transferred to other area of the cell to supply the membrane amplification.MreB works as the skeleton of bacteria,and in order to elucidate the mechanism of PlsB motility,we co-expressed EGFP-PlsB and Mchery-MreB in E.coli.We analyzed the co-localization microscope images of these two proteins,which showed EGFP-PlsB had similar localization pattern with that of mCherry-MreB,without significant co-localization.It comes to a conclusion that MreB was not a direct regulator for the movement of PlsB,but there might be a factor coupling mreB to PlsB.