Assessment of altered lipid homeostasis by HILIC-ion mobility-mass spectrometry-based lipidomics

Hines, Kelly M.; Herron, Josi; Xu, Libin

doi:10.1194/jlr.d074724

Cited by 90 publications

(123 citation statements)

References 64 publications

(95 reference statements)

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“…However, we found that total extractable lipids more than doubled (10.2%) when the cells were grown in the presence of 20% serum. Cells grown in the presence and absence of serum were subjected to comprehensive lipidomic analysis using hydrophilic interaction liquid chromatography-ion mobility-mass spectrometry (HILIC-IM-MS) (11, 29). The major lipid species observed in S. aureus grown in TSB included DGDGs, PGs, plasmalogen PGs (pPGs), and LysylPGs, as shown in the IM-extraction ion chromatogram (IM-XIC) in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Small aliquots were diluted into 2:1 acetonitrile/methanol for analysis. Extracts were analyzed by hydrophilic interaction chromatography (HILIC) coupled to an ion mobility-mass spectrometer (Synapt G2-Si IM-MS; Waters Corp., Milford, MA) in positive and negative modes (11, 29). Data analysis was performed with Progenesis QI (Nonlinear Dynamics; Waters Corp., Milford, MA) and lipid abundances were normalized to bacterial dry weight (11).…”

Section: Methodsmentioning

confidence: 99%

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Lipidomic and Ultrastructural Characterization of Cell Envelope ofStaphylococcus aureusGrown in the Presence of Human Serum

Hines

Alvarado

Chen

et al. 2020

Preprint

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26Staphylococcus aureus can incorporate exogenous straight-chain unsaturated and 27 saturated fatty acids (SCUFAs and SCFAs, respectively) to replace some of the normally 28 biosynthesized branched-chain fatty acids and SCFAs. In this study, the impact of human serum 29 on the S. aureus lipidome and cell envelope structure was comprehensively characterized. When 30 grown in the presence of 20% human serum, typical human serum lipids, such as cholesterol, 31 sphingomyelin, phosphatidylethanolamines, and phosphatidylcholines, were present in the total 32 lipid extracts. Mass spectrometry showed that SCUFAs were incorporated into all major S. 33 aureus lipid classes, i.e., phosphatidylglycerols, lysyl-phosphatidylglycerols, cardiolipins, and 34 diglucosyldiacylglycerols. Heat-killed S. aureus retained much fewer serum lipids and failed to 35 incorporate SCUFAs, suggesting that association and incorporation of serum lipids with S. 36 aureus requires a living or non-denatured cell. Cytoplasmic membranes isolated from 37 lysostaphin-produced protoplasts of serum-grown cells retained serum lipids, but washing cells 38 with Triton X-100 removed most of them. Furthermore, electron microscopy studies showed that 39 serum-grown cells had thicker cell envelopes and associated material on the surface, which was 40 partially removed by Triton X-100 washing. To investigate which serum lipids were 41 preferentially hydrolyzed by S. aureus lipases for incorporation, we incubated individual serum 42 lipid classes with S. aureus and found that cholesteryl esters (CEs) and triglycerides (TGs) are 43 the major donors of the incorporated fatty acids. Further experiments using purified Geh lipase 44 confirmed CEs and TGs being the substrates of this enzyme. Thus, growth in the presence of 45 serum altered the nature of the cell surface with implications for interactions with the host. 46 Page 3 of 38 IMPORTANCE 47Comprehensive lipidomics of S. aureus grown in the presence of human serum suggests 48 human serum lipids can associate with the cell envelope without being truly integrated into the 49 lipid membrane. However, fatty acids-derived from human serum lipids, including unsaturated 50 fatty acids, can be incorporated into lipid classes that can be biosynthesized by S. aureus itself. 51Cholesteryl esters and triglycerides are found to be the major source of incorporated fatty acids 52 upon hydrolysis by lipases. These findings have significant implications for the nature of the S. 53 aureus cell surface when grown in vivo. Changes in phospholipid and glycolipid abundances and 54 fatty acid composition could affect membrane biophysics and function and the activity of 55 membrane-targeting antimicrobials. Finally, the association of serum lipids with the cell envelope 56 has implications for the physicochemical nature of the cell surface and its interaction with host 57 defense systems. 58 KEYWORDS 59 Lipidomics, human serum lipids, fatty acid incorporation, lipid association, cell envelope 60 structure 61 62 63 64 65 6...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Lipidomic and Ultrastructural Characterization of Cell Envelope ofStaphylococcus aureusGrown in the Presence of Human Serum

Hines

Alvarado

Chen

et al. 2020

Preprint

Self Cite

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“…More recently, Rashid et al used a combination of targeted and untargeted LC-MS methods for comprehensive lipidomic studies of E. faecalis , but the workflow is time-consuming as separate sample preparations and LC-MS methods are needed for glycerolipids, phospholipids, and CLs ( 37 ). To address the limitations of the previous methods, we recently developed a 3D lipidomics approach based on hydrophilic interaction LC-ion mobility-MS (HILIC-IM-MS) that enables high-throughput and high-specificity lipid analysis ( 38 ). HILIC separates lipid classes on the basis of their head group polarity (on a scale of seconds), and IM separates lipid ions on the basis of their gas phase structures or collision cross sections (CCSs) as they pass through an inert gas background (on a scale of milliseconds).…”

Section: Introductionmentioning

confidence: 99%

Characterization of the Mechanisms of Daptomycin Resistance among Gram-Positive Bacterial Pathogens by Multidimensional Lipidomics

Hines

Waalkes

Penewit

et al. 2017

mSphere

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This work comprehensively characterizes lipidomic changes underlying daptomycin resistance in three Gram-positive bacterial species, E. faecalis, S. aureus, and C. striatum, by using a novel three-dimensional lipidomics methodology based on advanced mass spectrometry. We demonstrated a number of advantages of our method in comparison with other methods commonly used in the field, such as high molecular specificity, sensitivity, and throughput. Whole-genome sequencing of the S. aureus and C. striatum strains identified mutations in pgsA, which encodes phosphatidylglycerophosphate synthase, in both resistant strains. Lipidomics revealed significantly decreased levels of lipids downstream of PgsA, as well as accumulation of lipids upstream of PgsA in the resistant strains. Furthermore, we found that changes in individual molecular species of each lipid class depend on the their specific fatty acid compositions. The characteristic changes in individual lipid species could be used as biomarkers for identifying underlying resistance mechanisms and for evaluating potential therapies.

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“… 7 , 8 Moreover, even subclasses of each group of molecules, such as those of lipids, can be differentiated based on their location in the 2D spectrum. 8 − 12 Thus, IM-MS could be used to approximately classify an unknown ion based on its location in the CCS–mass 2D spectrum.…”

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Large-Scale Structural Characterization of Drug and Drug-Like Compounds by High-Throughput Ion Mobility-Mass Spectrometry

et al. 2017

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Ion mobility-mass spectrometry (IM-MS) can provide orthogonal information, i.e., m/z and collision cross section (CCS), for the identification of drugs and drug metabolites. However, only a small number of CCS values are available for drugs, which limits the use of CCS as an identification parameter and the assessment of structure–function relationships of drugs using IM-MS. Here, we report the development of a rapid workflow for the measurement of CCS values of a large number of drug or drug-like molecules in nitrogen on the widely available traveling wave IM-MS (TWIM-MS) platform. Using a combination of small molecule and polypeptide CCS calibrants, we successfully determined the nitrogen CCS values of 1425 drug or drug-like molecules in the MicroSource Discovery Systems’ Spectrum Collection using flow injection analysis of 384-well plates. Software was developed to streamline data extraction, processing, and calibration. We found that the overall drug collection covers a wide CCS range for the same mass, suggesting a large structural diversity of these drugs. However, individual drug classes appear to occupy a narrow and unique space in the CCS–mass 2D spectrum, suggesting a tight structure–function relationship for each class of drugs with a specific target. We observed bimodal distributions for several antibiotic species due to multiple protomers, including the known fluoroquinolone protomers and the new finding of cephalosporin protomers. Lastly, we demonstrated the utility of the high-throughput method and drug CCS database by quickly and confidently confirming the active component in a pharmaceutical product.

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Assessment of altered lipid homeostasis by HILIC-ion mobility-mass spectrometry-based lipidomics

Cited by 90 publications

References 64 publications

Lipidomic and Ultrastructural Characterization of Cell Envelope ofStaphylococcus aureusGrown in the Presence of Human Serum

Lipidomic and Ultrastructural Characterization of Cell Envelope ofStaphylococcus aureusGrown in the Presence of Human Serum

Characterization of the Mechanisms of Daptomycin Resistance among Gram-Positive Bacterial Pathogens by Multidimensional Lipidomics

Large-Scale Structural Characterization of Drug and Drug-Like Compounds by High-Throughput Ion Mobility-Mass Spectrometry

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