Tessa Poppenga’s research while affiliated with Ruhr University Bochum and other places

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Publications (1)


Fig 1. Lipid profiles of A. tumefaciens grown in the presence of different L-PLs. A. 2D-TLC profiles of selected lyso-PLs and PLs as references (standard) and A. tumefaciens grown in the absence (A. tumefaciens) or presence of 18:1 lyso-phosphatidylcholine (L-PC), 18:1 lyso-phosphatidylethanolamine (L-PE) and 18:1 L-PG (5 mM each) in LB medium. Lipids were visualized by CuSO 4 treatment. Arrows indicate accumulation of PC and PE in lipid spectra from L-PC-and L-PE-grown cultures. Two unknown lipids (named U1 and U2) were present in the lipid spectra of L-PG-grown cultures that accounted for 16.9% and 16.3% of total lipids, respectively (as quantified based on relative spot intensities from three independent experiments). 1D, first dimension; 2D, second dimension;?, unidentified lipid; PE, phosphatidylethanolamine; MMPE, monomethyl-PE; PG, phosphatidylglycerol; CL, cardiolipin; OL, ornithine lipids; PC, phosphatidylcholine. B. Structures of the lipid substrate L-PG and putative acylation products. BMP, bis(monoacylglycero)phosphate; R, alkyl group. [Color figure can be viewed at wileyonlinelibrary.com]
Fig 2. Identification of U1 and U2 as bis(monoacylglycero)phosphate. A. Q-TOF MS/MS spectra of U1 and U2 and of the reference lipids PG (18:1/18:1) and L-PG (18:0), analysed in the positive ion mode. PG (18:1/18:1) is detected as an ammonium adduct with m/z 792.5861 (parent ion). The predominating fragment ion with m/z 603.5431 is derived from diacylglycerol (DAG) with two 18:1 fatty acids (DAG-36:2). The neutral loss of 189.0430 u (= m/z 792.5861 À 603.5431) results from the glycerophosphate head group with ammonium. The minor fragment with m/z 339.2936 represents monoacylglycerol (MAG) with 18:1 fatty acids (MAG-18:1). L-PG (18:0) is detected as its protonated form with m/z 513.3258. The fragment ion with m/z 341.3098 represents MAG-18:0, and the neutral loss of 172.0160 u (= m/z 513.3258 minus 341.3098) is derived from glycerophosphate. U1 and U2 can be detected as ammonium adducts as shown for PG and with identical m/z values (792.584). However, their fragmentation patterns show only one fragment ion each with m/z 339.2969 (U1) or 339.2971 (U1), derived from MAG-18:1. Consequently, U1 and U2 do not contain DAG, but MAG units. B. Proposed structures for the fragment ions of PG (18:1/18:1), L-PG (18:0) and U1 or U2 (18:1/18:1). Masses as detected in MS/MS analysis are indicated. Note that fragmentation of U1/U2 results in masses corresponding to MAG. [Color figure can be viewed at wileyonlinelibrary.com]
Fig 3. Q-TOF MS/MS spectra of U1 and U2, PG and L-PG in the negative mode. A. In negative mode, PG (18:1/18:1) and U1/U2 are indistinguishable with similar fragmentation patterns and m/z values. L-PG (18:0) is detected in its deprotonated form with an m/z of 511.2911. B. Spiking of commercial BMP (18:1/18:1) into lipid extracts from cells grown in the presence and absence of L-PG (5 mM) establishes two isoforms of BMP (BMP-1 and BMP-2) as reaction products from L-PG. Lipids were visualized by Molybdenum Blue reagent. [Color figure can be viewed at wileyonlinelibrary.com]
Fig 4. Lipid profile of A. tumefaciens grown in the presence of putative substrates for BMP synthesis. A. 2D-TLC profiles revealed the absence of BMP-1 and BMP-2 from strains grown in the presence of 18:0/18:1 phosphatidylglycerol (+PG), glycerol 3-phosphate (+ G3P), 18:1 monoacylglycerol (+ MAG) or glycerol (+ Gly) (5 mM each) as visualized by CuSO 4 treatment. B. 2D-TLC of A. tumefaciens grown in the presence of different L-PG species suggests the production of BMP from both 18:1 and 18:0 L-PG as substrates. Lipids were visualized by Molybdenum Blue reagent. C. Strains lacking cardiolipin (Δcls1/Δcls2) or methylated PEderivatives MMPE and PC (ΔpmtA/Δpcs) still produce BMP-1 and BMP-2 when grown in the presence of (18:1) L-PG. Absence of CL, MMPE and PC in the respective mutant strains is indicated by dashed arrows. Lipids were visualized by CuSO 4 treatment. [Color figure can be viewed at wileyonlinelibrary.com]
Fig 5. L-PG acylation activity resides in membrane-derived fractions A. tumefaciens cultures were disrupted via ultrasonic treatment (crude extract) and separated from unbroken cells and debris to yield cell-free extracts. Ultracentrifugation allowed for the further separation into cytosolic and membrane fractions. After the addition of L-PG to each sample, total lipids were isolated after 3 h and 18 h of incubation at 30 C and monitored for BMP production via 1D-TLC. Arrows indicate the absence of BMP production from L-PG in the cytosolic samples (dashed line) or the production of BMP in membrane-derived samples (solid line). Lipids were visualized by CuSO 4 treatment.?: unidentified lipids. [Color figure can be viewed at wileyonlinelibrary.com]

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Synthesis of the unusual lipid bis(monoacylglycero)phosphate in environmental bacteria
  • Article
  • Full-text available

September 2021

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179 Reads

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3 Citations

Environmental Microbiology

Simon Czolkoss

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Pia Borgert

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Tessa Poppenga

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[...]

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The bacterial membrane is constantly remodelled in response to environmental conditions and the external supply of precursor molecules. Some bacteria are able to acquire exogenous lyso‐phospholipids and convert them to the corresponding phospholipids. Here we report that some soil‐dwelling bacteria have alternative options to metabolize lyso‐phosphatidylglycerol (L‐PG). We find that the plant‐pathogen Agrobacterium tumefaciens takes up this mono‐acylated phospholipid and converts it to two distinct isoforms of the non‐canonical lipid bis(monoacylglycero)phosphate (BMP). Chromatographic separation and Q‐TOF MS/MS analysis revealed the presence of two possible BMP stereo configurations acylated at either of the free hydroxyl groups of the glycerol head group. BMP accumulated in the inner membrane and did not visibly alter cell morphology and growth behaviour. The plant‐associated bacterium Sinorhizobium meliloti was also able to convert externally provided L‐PG to BMP. Other bacteria like Pseudomonas fluorescens and Escherichia coli metabolized L‐PG after cell disruption, suggesting that BMP production in the natural habitat relies both on dedicated uptake systems and on head‐group acylation enzymes. Overall, our study adds two previously overlooked phospholipids to the repertoire of bacterial membrane lipids and provides evidence for the remarkable condition‐responsive adaptation of bacterial membranes. This article is protected by copyright. All rights reserved.

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