Elucidation and identification of amino acid containing membrane lipids using liquid chromatography/high‐resolution mass spectrometry

Moore, Elizabeth Hernández; Hopmans, Ellen C.; Rijpstra, W. Irene C.; Villanueva, Laura; Damsté, Jaap S. Sinninghe

doi:10.1002/rcm.7503

Cited by 28 publications

(22 citation statements)

References 43 publications

(121 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The acylated amino acids GL and flavolipin have previously been identified in the membranes of several different members of the phylum Bacteroidetes (8,21,32). In this study, we have identified the genes required for the production of these acylated amino acids.…”

Section: Discussionmentioning

confidence: 92%

The Glycine Lipids of Bacteroides thetaiotaomicron Are Important for Fitness during Growth In Vivo and In Vitro

Lynch

Tammireddy

Doherty

et al. 2019

Appl Environ Microbiol

View full text Add to dashboard Cite

Acylated amino acids function as important components of the cellular membrane in some bacteria. Biosynthesis is initiated by the N-acylation of the amino acid, and this is followed by subsequent O-acylation of the acylated molecule, resulting in the production of the mature diacylated amino acid lipid. In this study, we use both genetics and liquid chromatography-mass spectrometry (LC-MS) to characterize the biosynthesis and function of a diacylated glycine lipid (GL) species produced in Bacteroides thetaiotaomicron. We, and others, have previously reported the identification of a gene, named glsB in this study, that encodes an N-acyltransferase activity responsible for the production of a monoacylated glycine called N-acyl-3-hydroxy-palmitoyl glycine (or commendamide). In all of the Bacteroidales genomes sequenced so far, the glsB gene is located immediately downstream from a gene, named glsA, that is also predicted to encode a protein with acyltransferase activity. We use LC-MS to show that the coexpression of glsB and glsA results in the production of GL in Escherichia coli. We constructed a deletion mutant of the glsB gene in B. thetaiotaomicron, and we confirm that glsB is required for the production of GL in B. thetaiotaomicron. Moreover, we show that glsB is important for the ability of B. thetaiotaomicron to adapt to stress and colonize the mammalian gut. Therefore, this report describes the genetic requirements for the biosynthesis of GL, a diacylated amino acid species that contributes to fitness in the human gut bacterium B. thetaiotaomicron. IMPORTANCE The gut microbiome has an important role in both health and disease of the host. The mammalian gut microbiome is often dominated by bacteria from the Bacteroidales, an order that includes Bacteroides and Prevotella. In this study, we have identified an acylated amino acid, called glycine lipid, produced by Bacteroides thetaiotaomicron, a beneficial bacterium originally isolated from the human gut. In addition to identifying the genes required for the production of glycine lipids, we show that glycine lipids have an important role during the adaptation of B. thetaiotaomicron to a number of environmental stresses, including exposure to either bile or air. We also show that glycine lipids are important for the normal colonization of the murine gut by B. thetaiotaomicron. This work identifies glycine lipids as an important fitness determinant in B. thetaiotaomicron and therefore increases our understanding of the molecular mechanisms underpinning colonization of the mammalian gut by beneficial bacteria.

show abstract

Section: Discussionmentioning

confidence: 92%

The Glycine Lipids of Bacteroides thetaiotaomicron Are Important for Fitness during Growth In Vivo and In Vitro

Lynch

Tammireddy

Doherty

et al. 2019

Appl Environ Microbiol

View full text Add to dashboard Cite

show abstract

“…The acylated amino acids, GL and flavolipin, have previously been identified in the membranes of several different members of the Phylum Bacteroidetes (8, 17, 32). In this study we have identified, for the first time, the genes required for the production of these acylated amino acids.…”

Section: Discussionmentioning

confidence: 99%

Characterization of the role of glycine lipids inBacteroides thetaiotaomicron

Lynch

Tammireddy

Doherty

et al. 2018

Preprint

View full text Add to dashboard Cite

18Acylated amino acids function as important components of the cellular membrane in some 19 bacteria. Biosynthesis is initiated by the N-acylation of the amino acid and this is followed by 20 subsequent O-acylation of the acylated molecule resulting in the production of the mature 21 diacylated amino acid lipid. In this study we use both genetics and liquid chromatography- 22 mass spectrometry (LC-MS) to characterize the biosynthesis and function of novel diacylated 23 glycine lipid (GL) species in Bacteroides thetaiotaomicron. We, and others, have previously 24 reported the identification of a gene, named glsB in this study, that encodes a N- 25 acyltransferase activity responsible for the production of a monoacylated glycine called N-26 acyl-3-hydroxy-palmitoyl glycine (or commendamide). In all of the Bacteroidales genomes so 27 far sequenced the glsB gene is located immediately downstream from a gene, named glsA, 28 also predicted to encode a protein with acyltransferase activity. We use LC-MS to show that 29 co-expression of glsB and glsA results in the production of GL in Escherichia coli. We 30 constructed a deletion mutant of the glsB gene in B. thetaiotaomicron and we confirm that 31 glsB is required for the production of GL in B. thetaiotaomicron. Moreover, we show that glsB 32 is important for the ability of B. thetaiotaomicron to adapt to stress and colonize the 33 mammalian gut. Therefore, this report is the first to describe the genetic requirements for 34 the biosynthesis of GL, a novel diacylated amino acids species that contributes to fitness in 35 the human gut bacterium, B. thetaiotaomicron. 36 37 38 39 42 healthy human gut microbiota (1). The gut-associated Bacteroidetes are required to digest 43 complex dietary glycans into short-chain fatty acids (such as acetate and propionate) that 44 are accessible to the host (2-4). A longitudinal study in infants has revealed the presence of 45 Bacteroides in the infant gut within 1 week of birth and some species of Bacteroides have 46 been shown to utilize the polysaccharides present in human breast milk (5, 6). Therefore it 47 has been suggested that Bacteroides may have an important role during the early 48 development of the infant gut (6). 49 50Acylated amino acids can be found in the membranes of many bacteria (7, 8). The best-51 characterized, and most widespread, acylated amino acid is ornithine lipid (OL). OL contains 52 a 3'-hydroxy fatty acid group attached by an amide linkage to the a-amino group of 53 ornithine with a second fatty acid group ester linked to the 3'-hydroxy group of the first 54 fatty acid (9). The genetics of OL production was first described in Sinorhizobium meliloti 55 where it was shown that a N-acyltransferase encoded by olsB catalysed the attachment of 56 the first fatty acid group to ornithine, resulting in monoacylated ornithine or lyso-OL (10). 57 The second fatty acid was subsequently attached to lyso-OL through the activity of an O-58 acyltransferase encoded by olsA, resulting in the producti...

show abstract

“…The dried extract residue was dissolved in hexane/2-propanol/H 2 O (718:271:10, v/v/v) injection solvent, filtered through a 0.45 µm, 4 mm diameter True TM Regenerated Cellulose syringe filter (Grace Davison) prior to injection and analysis. Extracted IPLs from the enrichment slurries were analyzed by high-performance liquid chromatography-electrospray ionization-ion trap mass spectrometry (HPLC-ESI/IT/MS) according to [34], with some modifications [35]. IPL abundance was assessed by integrating the HPLC-ESI/IT/MS mass chromatogram for each individual identified IPL, and areas were summed per IPL head group and reported per gram of slurry, dry weight.…”

Section: Intact Polar Lipid (Ipl) Extraction and Hplc-esi/it/ms Analysismentioning

confidence: 99%

Butyrate Conversion by Sulfate-Reducing and Methanogenic Communities from Anoxic Sediments of Aarhus Bay, Denmark

et al. 2020

Self Cite

View full text Add to dashboard Cite

The conventional perception that the zone of sulfate reduction and methanogenesis are separated in high- and low-sulfate-containing marine sediments has recently been changed by studies demonstrating their co-occurrence in sediments. The presence of methanogens was linked to the presence of substrates that are not used by sulfate reducers. In the current study, we hypothesized that both groups can co-exist, consuming common substrates (H2 and/or acetate) in sediments. We enriched butyrate-degrading communities in sediment slurries originating from the sulfate, sulfate–methane transition, and methane zone of Aarhus Bay, Denmark. Sulfate was added at different concentrations (0, 3, 20 mM), and the slurries were incubated at 10 °C and 25 °C. During butyrate conversion, sulfate reduction and methanogenesis occurred simultaneously. The syntrophic butyrate degrader Syntrophomonas was enriched both in sulfate-amended and in sulfate-free slurries, indicating the occurrence of syntrophic conversions at both conditions. Archaeal community analysis revealed a dominance of Methanomicrobiaceae. The acetoclastic Methanosaetaceae reached high relative abundance in the absence of sulfate, while presence of acetoclastic Methanosarcinaceae was independent of the sulfate concentration, temperature, and the initial zone of the sediment. This study shows that there is no vertical separation of sulfate reducers, syntrophs, and methanogens in the sediment and that they all participate in the conversion of butyrate.

show abstract

Elucidation and identification of amino acid containing membrane lipids using liquid chromatography/high‐resolution mass spectrometry

Cited by 28 publications

References 43 publications

The Glycine Lipids of Bacteroides thetaiotaomicron Are Important for Fitness during Growth In Vivo and In Vitro

The Glycine Lipids of Bacteroides thetaiotaomicron Are Important for Fitness during Growth In Vivo and In Vitro

Characterization of the role of glycine lipids inBacteroides thetaiotaomicron

Butyrate Conversion by Sulfate-Reducing and Methanogenic Communities from Anoxic Sediments of Aarhus Bay, Denmark

Contact Info

Product

Resources

About