An advanced method for propargylcholine phospholipid detection by direct-infusion MS

Abstract

Phospholipids with a choline head group are an abundant component of cellular membranes and are involved in many important biological functions. For studies on the cell biology and metabolism of these lipids, traceable analogues where propargylcholine replaces the choline head group have proven useful. We present a novel method to analyze propargylcholine phospholipids by MS. The routine employs 1-radyl-2-lyso-sn-glycero-3-phosphopropargylcholines as labeled lysophosphatidylcholine precursors, which upon cellular conversion direct the traceable tag with superb specificity and efficiency to the primary target lipid class. Using azidopalmitate as a click-chemistry reporter, we introduce a highly specific, sensitive, and robust MS detection procedure for the propargylcholine phospholipids. In a first study, we apply the new technique to investigate choline phospholipid metabolism in brain endothelial cells. These experiments reveal differences in the metabolism of phosphatidylcholine and its pendant, ether phosphatidylcholine. The novel method described here opens a new, quantitative, and detailed view on propargylcholine phospholipid metabolism and will greatly facilitate future studies on choline phospholipid metabolism.

The foundations and development of lipidomics

“…These include both the use of multistage fragmentation to identify sn -1 versus sn -2 chains, location of double bond(s) in fatty acyl-chains, and cis - trans isomerism. To this end, newly developed technology/instrumentation including ultraviolet photodissociation MS ( 175 ) and a variety of chemical reactions/derivatization approaches have been broadly used for deep analysis of lipid molecular structures, including Paternò–Büchi reaction ( 176 , 177 , 178 ) and click chemistry ( 179 , 180 , 181 ). Readers interested in this area of work can find the comprehensive review of deep phenotyping with chemical reaction/derivatization approaches by Xia in this Thematic Review series.…”

Tracing Lipid Metabolism by Alkyne Lipids and Mass Spectrometry: The State of the Art

“…The water-soluble alcohol was administered to the biological specimen and entered lipid metabolism via the Kennedy pathway or PLD remodeling ( Bumpus et al, 2018 ). Alternatively, a labeling strategy using synthetic lyso-phosphatidylpropargylcholine LpPC or its ether pendant LpPC O was used to specifically target the PC or PC O pool, respectively ( Yaghmour et al, 2021 ).…”

“…However, this approach only delivered their sum FA composition. This limitation has recently been overcome with the introduction of a novel azide reporter, azidopalmitate ( Figure 6A ), for propargylcholine-containing phospholipids ( Yaghmour et al, 2021 ).…”

“…The new method was used to investigate the cellular PC and PC O content. The study revealed differences in pool size, apparent metabolic stability, and side-chain composition of both lipid classes by providing a quantitative picture of their metabolism and homeostasis ( Yaghmour et al, 2021 ).…”

Development of oxaalkyne and alkyne fatty acids as novel tracers to study fatty acid beta-oxidation pathways and intermediates

“…Clickable lipid tracers were successfully used to monitor protein lipidation, protein-lipid interaction, lipid localization, and metabolism, and this work has been extensively reviewed ( 16 , 17 , 18 , 19 , 20 ). Importantly, alkyne lipids are amenable to various highly sensitive detection procedures including TLC and MS-based lipidomics ( 11 , 21 , 22 , 23 , 24 , 25 , 26 , 27 ). The versatility of the alkyne tag in combination with advanced click reporters, detection technologies, and instrumentation has opened new possibilities in lipid research.…”