A study of membrane lipids from dehydration-acclimated Brassica napus root cells: formation of a cubic phase under physiological conditions

1990 ◽  
Vol 68 (1) ◽  
pp. 102-105 ◽  
Author(s):  
P. Norberg ◽  
K. Larsson ◽  
C. Liljenberg

Rape seedlings were acclimated to evaporative dehydration by exposure to repeated moderate water-deficit stress. The stress program started after 19 days of growth and consisted of three, 24-h stress periods interspersed with 24-h rewatering periods. After the third stress period the roots were harvested and microsomal membranes were isolated. Control plants were grown under equivalent conditions without stress (nonacclimated cells). Total lipids were extracted from the membranes and investigated with X-ray crystallography and polarization microscopy at different degrees of hydration and temperatures. In excess water, the membrane lipids from both acclimated and nonacclimated cells exhibited a cubic phase. The lipids from the nonacclimated cells formed a hexagonal (HII) phase on dehydration. The lipids from the acclimated cells behaved in a different way during dehydration, where the cubic phase was transformed to an L2 phase via an intermediate HII phase. At increasing temperatures, the hydrated cubic phase started to form an L2 phase at 30 °C and was fully converted to the liquid-type state at 42 °C. The mesomorphic phase behaviour is discussed in relation to membrane activity.Key words: water-deficit stress, microsomal membranes, X-ray crystallography, polarization microscopy.

2001 ◽  
Vol 26 (2) ◽  
pp. 106-112 ◽  
Author(s):  
Paul K Fyfe ◽  
Katherine E McAuley ◽  
Aleksander W Roszak ◽  
Neil W Isaacs ◽  
Richard J Cogdell ◽  
...  

1985 ◽  
Vol 63 (2) ◽  
pp. 77-84 ◽  
Author(s):  
C. Liljenberg ◽  
M. Kates

The effect of repeated water-deficit stress on the lipid composition of root cells from 5-day-old oat (Avena sativa L. cv. Seger) seedlings was studied. The content of total acyl lipids was found to decrease with increasing degree of water-deficit stress, owing largely to decreases in free fatty acids, triglycerides, phosphatidylethanolamine (PE), wax esters, steryl esters, and acylated steryl glycosides. Major polar lipids both in total root cells and in the plasma membrane enriched fraction, as well as the microsomal membrane fraction, were PE, phosphatidylcholine (PC), digalactosyldiacylglycerol (DGDG), monogalactosyldiacylglycerol (MGDG), and polyglycolipid. Decreases in the degree of unsaturation of the fatty acids as a funtion of increased water-deficit stress were observed for the MGDG and polyglycolipid components of total root cells and for the MGDG, DGDG, and polyglycolipid of the plasma membrane fraction. Electron microscopy showed that stressed root tip cells had much smoother plasma membranes than those of control unstressed root cells. These results suggest that root cells of oat seedlings respond to water-deficit stress by reducing the total plasma membrane mass and degree of lipid fluidity, which would reduce the water permeability of the plasma membranes and help maintain cell turgidity.


2017 ◽  
Vol 24 (5) ◽  
pp. 1086-1091 ◽  
Author(s):  
Minoru Kubo ◽  
Eriko Nango ◽  
Kensuke Tono ◽  
Tetsunari Kimura ◽  
Shigeki Owada ◽  
...  

X-ray free-electron lasers (XFELs) have opened new opportunities for time-resolved X-ray crystallography. Here a nanosecond optical-pump XFEL-probe device developed for time-resolved serial femtosecond crystallography (TR-SFX) studies of photo-induced reactions in proteins at the SPring-8 Angstrom Compact free-electron LAser (SACLA) is reported. The optical-fiber-based system is a good choice for a quick setup in a limited beam time and allows pump illumination from two directions to achieve high excitation efficiency of protein microcrystals. Two types of injectors are used: one for extruding highly viscous samples such as lipidic cubic phase (LCP) and the other for pulsed liquid droplets. Under standard sample flow conditions from the viscous-sample injector, delay times from nanoseconds to tens of milliseconds are accessible, typical time scales required to study large protein conformational changes. A first demonstration of a TR-SFX experiment on bacteriorhodopsin in bicelle using a setup with a droplet-type injector is also presented.


1985 ◽  
Vol 63 (3) ◽  
pp. 253-257 ◽  
Author(s):  
Conny Liljenberg ◽  
Pirjo Karunen ◽  
Rainer Ekman

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Jianping Li ◽  
Yang Shen ◽  
Yanke Chen ◽  
Zhengfeng Zhang ◽  
Shaojie Ma ◽  
...  

AbstractDiacylglycerol kinase (DgkA) is a small integral membrane protein, responsible for the ATP-dependent phosphorylation of diacylglycerol to phosphatidic acid. Its structures reported in previous studies, determined in detergent micelles by solution NMR and in monoolein cubic phase by X-ray crystallography, differ significantly. These differences point to the need to validate these detergent-based structures in phospholipid bilayers. Here, we present a well-defined homo-trimeric structure of DgkA in phospholipid bilayers determined by magic angle spinning solid-state NMR (ssNMR) spectroscopy, using an approach combining intra-, inter-molecular paramagnetic relaxation enhancement (PRE)-derived distance restraints and CS-Rosetta calculations. The DgkA structure determined in lipid bilayers is different from the solution NMR structure. In addition, although ssNMR structure of DgkA shows a global folding similar to that determined by X-ray, these two structures differ in monomeric symmetry and dynamics. A comparative analysis of DgkA structures determined in three different detergent/lipid environments provides a meaningful demonstration of the influence of membrane mimetic environments on the structure and dynamics of membrane proteins.


2020 ◽  
Author(s):  
Michael W. Martynowycz ◽  
Anna Shiriaeva ◽  
Xuanrui Ge ◽  
Johan Hattne ◽  
Brent L. Nannenga ◽  
...  

AbstractG Protein-Coupled Receptors (GPCRs), or 7-transmembrane receptors, are a superfamily of membrane proteins that are critically important to physiological processes in the human body. Determining high-resolution structures of GPCRs without signaling partners bound requires crystallization in lipidic cubic phase (LCP). GPCR crystals grown in LCP are often too small for traditional X-ray crystallography. These microcrystals are ideal for investigation by microcrystal electron diffraction (MicroED), but the gel-like nature of LCP makes traditional approaches to MicroED sample preparation insurmountable. Here we show that the structure of a human A2A adenosine receptor can be determined by MicroED after converting the LCP into the sponge phase followed by cryoFIB milling. We determined the structure of the A2A receptor to 2.8 Å resolution and resolved an antagonist in its orthosteric ligand-binding site as well as 4 cholesterol molecules bound to the receptor. This study lays the groundwork for future GPCR structural studies using single microcrystals that would otherwise be impossible by other crystallographic methods.One sentence summaryFIB milled LCP-GPCR structure determined by MicroED


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