Heterologous overexpression of a monotopic glucosyltransferase (MGS) induces fatty acid remodeling in Escherichia coli membranes

Abstract

The membrane protein monoglucosyldiacylglycerol synthase (MGS) from Acholeplasma laidlawii is responsible for the creation of intracellular membranes when overexpressed in Escherichia coli (E. coli). The present study investigates time dependent changes in composition and properties of E. coli membranes during 22h of MGS induction. The lipid/protein ratio increased by 38% in MGS-expressing cells compared to control cells. Time-dependent screening of lipids during this period indicated differences in fatty acid modeling. (1) Unsaturation levels remained constant for MGS cells (~62%) but significantly decreased in control cells (from 61% to 36%). (2) Cyclopropanated fatty acid content was lower in MGS producing cells while control cells had an increased cyclopropanation activity. Among all lipids, phosphatidylethanolamine (PE) was detected to be the most affected species in terms of cyclopropanation. Higher levels of unsaturation, lowered cyclopropanation levels and decreased transcription of the gene for cyclopropane fatty acid synthase (CFA) all indicate the tendency of the MGS protein to force E. coli membranes to alter its usual fatty acid composition.

Protein Amphipathic Helix Insertion: A Mechanism to Induce Membrane Fission

“…Class II: Passive mechanism, normal topology. Examples: fission mediated, in the absence of nucleoside triphosphate hydrolysis, by Bacillus subtilis FisB (Doan et al, 2013), C-terminal Binding Protein 1 Short form/Brefeldin A ADP-Ribosylation substrate (CtBP1-S/BARS) (Spanò et al, 1999;Weigert et al, 1999;Hidalgo Carcedo et al, 2004;Valente et al, 2005Valente et al, , 2012Liberali et al, 2008;Pagliuso et al, 2016;Zhukovsky et al, 2019b), and by numerous AH-containing proteins, such as endophilins (Rostovtseva et al, 2009;Ambroso et al, 2014;Genet et al, 2019), amphiphysins (Wu and Baumgart, 2014;Snead et al, 2019), epsins (Ford et al, 2002;Boucrot et al, 2012;Brooks et al, 2015), α-synuclein (Nakamura et al, 2011;Braun et al, 2017;Pozo Devoto and Falzone, 2017;Fakhree et al, 2019), GDAP1 (Huber et al, 2016), ankyrin repeats and KH domain-containing protein 1 (ANKHD1) (Kitamata et al, 2019), Saccharomyces cerevisiae Atg18 (Gopaldass et al, 2017), Agrobacterium tumefaciens PmtA (Danne et al, 2017b), EcMurG (van den Brink-van der Laan et al, 2003, Acholeplasma laidlawii MGS (Eriksson et al, 2009;Ariöz et al, 2014;Ge et al, 2014) and DGS (Eriksson et al, 2009).…”

Characterization of extracellular vesicles by IR spectroscopy: Fast and simple classification based on amide and C H stretching vibrations

“…5 l of sample was mounted on the diamond ATR crystal and a thin dry film was obtained by slowly evaporation of the buffer solvent under ambient conditions [33]. The measurements were performed at room temperature, immediately after drying the sample (within approximately 5 minutes).…”

“…We have to point out, however, that since the absorption coefficients of the protein-and lipid-related bands are different, for real biological samples without a proper calibration procedure only a 'spectroscopic' protein-to-lipid ratio can be determined [32]. The 'spectroscopic' protein-tolipid ratio, however, has been successfully applied in realistic measures of differences in protein-to-lipid ratios from one membrane to another investigating thylakoid and cytoplasmic membranes of cyanobacteria [32] or Escherichia coli membranes [33].…”

Metabolic phospholipid labeling of intact bacteria enables a fluorescence assay that detects compromised outer membranes

“…All PLs (PG, PE, PC, PPC and AEPC) were scanned for the 11 most abundant acyl chain combinations (Table S3) (33,48). Analyst® Software (SCIEX) was used for data acquisition.…”

“…2). In all BW25113 pcs samples grown with Cho, PC was observed in addition to PE and PG ( Table S3) (33,48). The isobaric ions of PC with equal sums of the two acyl chain lengths and number of double bonds (e.g.…”