Identification of phosphatidylserylglutamate: a novel minor lipid in Escherichia coli

Abstract

dylglycerol phosphate ( 3 ). For each of these lipids, there can exist several molecular species arising from the different lengths, unsaturation, and/or cyclopropane analogs of the acyl chains ( 4 ). The complexity of the E. coli lipidome, however, is even greater than can be explained by acyl chain heterogeneity. Numerous additional minor lipids are present in wild-type cells, as judged by isotopic labeling experiments and two-dimensional TLC ( 3 ). Many of these species cannot be identifi ed by their migration with standards of known lipids or biosynthetic precursors.Electrospray ionization-mass spectrometry (ESI-MS) is well suited to the analysis of intact lipids ( 5 ). Fragmentation during ionization is minimized and the sensitivity is high ( 6, 7 ). In addition, collision-induced dissociation mass spectrometry (MS/MS) allows for the structural analysis of the lipid ion of interest and, when combined with the high mass accuracy of time-of-fl ight mass spectrometers, can be used to propose a molecular formula for a particular ion.Current applications of mass spectrometry to lipid analysis have focused mainly on the quantifi cation of known lipid species, often coupling liquid chromatography directly to the mass spectrometer ( 8-10 ). These analyses compare levels of known lipid species present under various growth conditions or disease states. However, important changes in levels of unknown or minor lipids are diffi cult to analyze without knowledge of their structures and the availability of appropriate standards. Phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and cardiolipin (CL) are the major membrane glycerophospholipids of Escherichia coli , constituting about 10% of the dry weight of the cell ( 1, 2 ). Important minor lipids include the precursors phosphatidic acid, CDP-diacylglycerol, phosphatidylserine, and phosphati-

Altered Lipid Synthesis by Lack of Yeast Pah1 Phosphatidate Phosphatase Reduces Chronological Life Span

“…The fatty acid composition of the mitochondrial lipids after deacylation was determined by gas-liquid chromatography (35) using heptadecanoic acid as an internal standard. The mitochondrial phospholipid molecular species were analyzed by liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (62,63). For radiolabeling of cellular neutral lipids and total phospholipids, cells were grown in SC medium to the late exponential phase (A 600 nm ϳ1).…”

Non-enzymatically derived minor lipids found in Escherichia coli lipid extracts

“…The cellular lipids were extracted using the method of Bligh and Dyer [8] as described previously [1]. Briefly, 50 mL of chloroform and 100 mL of methanol were added to the cell suspension to generate a single-phase extraction mixture of CHCl 3 /CH 3 OH/PBS (1:2:0.8, v/v/v).…”

“…High-resolution mass spectrometry is a superb tool for the identification of new metabolites especially lipid metabolites. The combination of accurate mass measurements and collision-induced dissociation MS/MS analysis facilitates the identification of known and novel lipid structures and has been used to characterize several new lipid metabolites in Gram-negative bacteria such as Escherichia coli and other organisms such as Clostridium [1-5]. …”

Putative N-Acylphosphatidylethanolamine Synthase from Arabidopsis thaliana Is a Lysoglycerophospholipid Acyltransferase

“…The cellular lipids were extracted using the method of Bligh and Dyer (23), as described previously (24). Briefly, 1 ml of CHCl 3 and 2 ml of CH 3 OH were added to the cell suspension to generate a single phase extraction mixture of CHCl 3 /CH 3 OH/PBS (1:2:0.8, v/v/v).…”

“…The normal phase chromatography was performed using a Zorbax Rx-SIL (5 m, 4.6 ϫ 250 mm) column on an Agilent 1100 HPLC system as described previously (24). The HPLC effluent (0.5 ml/min) was analyzed using an Agilent 6520 quadrupole time-of-flight mass spectrometer.…”