Lipodisqs for eukaryote lipidomics with retention of viability: Sensitivity and resistance to Leucobacter infection linked to C.elegans cuticle composition

Abstract

Lipodisq™ nanoparticles have been used to extract surface lipids from the cuticle of two strains (wild type, N2 and the bacterial-resistant strain, agmo-1) of the C. elegans nematode without loss of viability. The extracted lipid were characterized by thin layer chromatography and MALDI-TOF-MS. The lipid profiles differed between the two strains. The extracted lipids from the bacterial resistant strain, agmo-1, contained ether-linked (O-alkyl chain) lipids, in contrast to the wild type strain which contained exclusively ester (O-acyl) linked lipids. This observation is consistent with the loss of a functional alkylglycerol monooxygenase (AGMO) in the bacterial resistant strain agmo-1. The presence and abundance of other lipid species also differs between the wild-type N2 and agmo-1 nematodes, suggesting that the agmo-1 mutant strain attempts to compensate for the increase in ether-linked lipids by modulating other lipidsynthesis pathways. Together these differences not only affect the fragility of the cuticle and the buoyancy of the worm in aqueous buffer, but also interactions with surface-adhering bacteria. The much greater chemical stability of O-alkyl, hydrolysable linked lipids compared with non-hydrolysable O-acyl linked lipids, may be the origin of the bacterial resistance of the agmo-1 strain, providing a more resilient cuticle for the nematode. Additionally, we show that lipid extraction with a polymer of styrene and maleic acid (SMA) provides a viable route to lipidomics studies with minimal perturbation of the organism.

Detergent-free extraction of a functional low-expressing GPCR from a human cell line

“…Several membrane proteins including ion channels [40] and GPCRs [41,42] have been studied and characterized from different organisms [43][44][45][46]. Proteomics and lipidomics experiments using SMALPs have also been performed using hydrogen-deuterium exchange mass spectrometry, MALDI-TOF MS/MS and using LC-MS/MS [47][48][49][50].…”

From polymer chemistry to structural biology: The development of SMA and related amphipathic polymers for membrane protein extraction and solubilisation

“…3:1 SMA, when combined with phospholipids, forms discoidal nanostructures with a typical size of around 11 nm in diameter (Figure 2a). This property has been exploited for lipidomics studies [32][33][34] , in particular in eukaryotic systems in which lipid extraction does not result in a loss of organism viability (see, Bada Juarez et al, this volume 35 ). Aqueous suspensions of these Lipodisq nanoparticles (also known as SMALPs) are monodisperse 36,37 , however at high concentrations they may form stacks (rouleaux), especially when deposited onto surfaces (Figure 2b).…”

Metal‐Chelated Polymer Nanodiscs for NMR Studies

“…The introduction of amphiphilic polymers that form lipid nanodiscs has most notably contributed to the study of membrane proteins (MPs) and allows for their functional and structural study in a tunable native‐like membrane environment . While polymer nanodiscs are the youngest of the nanodisc field, they are showing great potential due to the simplicity of their synthesis, their diverse chemical tunability, and their ability to directly extract membrane proteins from their cellular environment at practical cost . Nanodiscs are a useful tool for NMR spectroscopy since their size can be tuned to conditions favorable to both solution and solid‐state NMR.…”