Abstract
Lipid membranes are the border between living cells and their environments. The membrane's lipid composition defines fluidity, thickness, and protein activity and is controlled by the intricate actions of lipid gene-encoded enzymes. However, a comprehensive analysis of each protein's contribution to the lipidome is lacking. Here, we present such a comprehensive and functional overview of lipid genes in Escherichia coli by individual overexpression or deletion of these genes. We developed a high-throughput lipidomic platform, combining growth analysis, one-step lipid extraction, rapid LC-MS, and bioinformatic analysis into one streamlined procedure. This allowed the processing of more than 300 samples per day and revealed interesting functions of known enzymes and distinct effects of individual proteins on the phospholipidome. Our data demonstrate the plasticity of the phospholipidome and unexpected relations between lipid classes and cell growth. Modeling of lipidomic responses to short-chain alcohols provides a rationale for targeted membrane engineering.
Highlights
Escherichia coli (E. coli) is a popular biological production platform, but the recombinant production of eukaryotic membrane proteins and achieving high titers of hydrophobic compounds remain challenging because of their interaction with membrane phospholipids (Atsumi et al, 2008; Baumgarten et al, 2017; Choi and Lee, 2013; Rau et al, 2016; Schlegel et al, 2014; Yang et al, 2018)
A High-Throughput Method for Lipidomic Analysis of E. coli Mutants By combining lipid-enzyme activities reported in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways (Kanehisa and Goto, 2000) with current literature, we constructed a schematic overview of lipid metabolism in E. coli in which the lipid enzymes were classified in five pathway groups (Figure 1; Table S1)
In order to study the effect of these E. coli lipid enzymes on the lipidome and bacterial growth rates, we developed a high-throughput lipidomic platform
Summary
Escherichia coli (E. coli) is a popular biological production platform, but the recombinant production of eukaryotic membrane proteins and achieving high titers of hydrophobic compounds remain challenging because of their interaction with membrane phospholipids (Atsumi et al, 2008; Baumgarten et al, 2017; Choi and Lee, 2013; Rau et al, 2016; Schlegel et al, 2014; Yang et al, 2018). Many membrane proteins require a specific lipid context for optimal function (Wikstrom et al, 2009). The selective barrier function of membranes can be compromised by the production of non-physiological (levels of) hydrophobic compounds (Sikkema et al, 1995). Many membrane proteins bind lipids selectively to modulate their structure and function (Laganowsky et al, 2014). A thorough understanding of the phospholipidome is a prerequisite for the optimal exploitation of the potential of E. coli as a biotechnological platform
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