Back Cover: Thermodriven Micrometer-Scale Aqueous-Phase Separation of Amphiphilic Oligoethylene Glycol Analogues (Chem. Asian J. 10/2014)

Treatment of kidney microvillar membranes with the non-ionic detergent Triton X-114 at 0 degrees C, followed by low-speed centrifugation, generated a detergent-insoluble pellet and a detergent-soluble supernatant. The supernatant was further fractionated by phase separation at 30 degrees C into a detergent-rich phase and a detergent-depleted or aqueous phase. Those ectoenzymes with a covalently attached glycosyl-phosphatidylinositol (G-PI) membrane anchor were recovered predominantly (greater than 73%) in the detergent-insoluble pellet. In contrast, those ectoenzymes anchored by a single membrane-spanning polypeptide were recovered predominantly (greater than 62%) in the detergent-rich phase. Removal of the hydrophobic membrane-anchoring domain from either class of ectoenzyme resulted in the proteins being recovered predominantly (greater than 70%) in the aqueous phase. This technique was also applied to other membrane types, including pig and human erythrocyte ghosts, where, in both cases, the G-PI-anchored acetylcholinesterase partitioned predominantly (greater than 69%) into the detergent-insoluble pellet. When the microvillar membranes were subjected only to differential solubilization with Triton X-114 at 0 degrees C, the G-PI-anchored ectoenzymes were recovered predominantly (greater than 63%) in the detergent-insoluble pellet, whereas the transmembrane-polypeptide-anchored ectoenzymes were recovered predominantly (greater than 95%) in the detergent-solubilized supernatant. Thus differential solubilization and temperature-induced phase separation in Triton X-114 distinguished between G-PI-anchored membrane proteins, transmembrane-polypeptide-anchored proteins and soluble, hydrophilic proteins. This technique may be more useful and reliable than susceptibility to release by phospholipases as a means of identifying a G-PI anchor on an unpurified membrane protein.

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The hyaluronate synthase from a eukaryotic cell line

Biochemical Journal ◽

10.1042/bj2900791 ◽

1993 ◽

Vol 290 (3) ◽

pp. 791-795 ◽

Cited By ~ 25

Author(s):

L Klewes ◽

E A Turley ◽

P Prehm

Keyword(s):

Monoclonal Antibodies ◽

Phase Separation ◽

Affinity Chromatography ◽

Cell Line ◽

Aqueous Phase ◽

Plasma Membranes ◽

Eukaryotic Cell ◽

Molecular Masses ◽

Triton X ◽

Hyaluronate Synthase

The hyaluronate synthase complex was identified in plasma membranes from B6 cells. It contained two subunits of molecular masses 52 kDa and 60 kDa which bound the precursor UDP-GlcA in digitonin solution and partitioned into the aqueous phase, together with nascent hyaluronate upon Triton X-114 phase separation. The 52 kDa protein cross-reacted with poly- and monoclonal antibodies raised against the streptococcal hyaluronate synthase and the 60 kDa protein was recognized by monoclonal antibodies raised against a hyaluronate receptor. The 52 kDa protein was purified to homogeneity by affinity chromatography with monoclonal anti-hyaluronate synthase.

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Aqueous Phase Separation: a novel and sustainable approach for membrane preparation

10.3990/1.9789036552103 ◽

2021 ◽

Author(s):

Wouter Nielen

Keyword(s):

Phase Separation ◽

Aqueous Phase ◽

Membrane Preparation

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Aqueous phase separation: A basic physicochemical phenomenon with a potential role in the evolution and organization of the liquid phase of cytoplasm

Origins of Life and Evolution of Biospheres ◽

10.1007/bf02459861 ◽

1996 ◽

Vol 26 (3-5) ◽

pp. 448-449

Author(s):

Harry Walter ◽

Donald E. Brooks

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Aqueous Phase ◽

Potential Role ◽

Physicochemical Phenomenon

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Powerful Concentration of Rhodium in Plating Wastewater Using Homogeneous Liquid–Liquid Extraction (HoLLE) and Study for Scale-up

Environment and Natural Resources Research ◽

10.5539/enrr.v7n4p44 ◽

2017 ◽

Vol 7 (4) ◽

pp. 44 ◽

Cited By ~ 1

Author(s):

Takeshi Kato ◽

Shotaro Saito ◽

Shigekatsu Oshite ◽

Shukuro Igarashi

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Aqueous Phase ◽

Scale Up ◽

Volume Ratio ◽

Liquid Extraction ◽

Optimum Conditions ◽

Homogeneous Liquid ◽

Liquid Liquid Extraction ◽

Plating Wastewater

A powerful technique for the concentration of rhodium (Rh) in plating wastewater was developed. The technique entails complexing Rh with 1-(2-pyridylazo)-2-naphthol (PAN) followed by homogeneous liquid–liquid extraction (HoLLE) with Zonyl FSA. The optimum HoLLE conditions were determined as follows: [ethanol]T = 30.0 vol.%, pH = 4.00, and Rh:PAN = 1:5. Under these optimum conditions, 88.1% of Rh was extracted into the sedimented liquid phase. After phase separation, the volume ratio [aqueous phase (Va) /sedimented liquid phase (Vs)] of Va and Vs was 1000 (50 mL → 0.050 mL). We then applied the new method to wastewater generated by the plating industry. The phase separation was satisfactorily achieved when the volume was scaled up to 1000 mL of the actual wastewater; 84.7% of Rh was extracted into the sedimented liquid phase. After phase separation, Va/Vs was 588 (1000 mL - 1.70 mL).

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Aqueous Phase Separation Behavior of Highly Syndiotactic, High Molecular Weight Polymers with Densely Packed Hydroxy-Containing Side Groups

Macromolecules ◽

10.1021/acs.macromol.8b01150 ◽

2018 ◽

Vol 51 (18) ◽

pp. 7248-7256 ◽

Cited By ~ 8

Author(s):

Dorette S. Tromp ◽

Marianne Lankelma ◽

Hannah de Valk ◽

Emile de Josselin de Jong ◽

Bas de Bruin

Keyword(s):

Molecular Weight ◽

Phase Separation ◽

High Molecular Weight ◽

Aqueous Phase ◽

High Molecular Weight Polymers ◽

Phase Separation Behavior ◽

Separation Behavior

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Rapid Multilevel Compartmentalization of Stable All-Aqueous Blastosomes by Interfacial Aqueous-Phase Separation

ACS Nano ◽

10.1021/acsnano.0c02923 ◽

2020 ◽

Vol 14 (9) ◽

pp. 11215-11224

Author(s):

Shipei Zhu ◽

Joe Forth ◽

Ganhua Xie ◽

Youchuang Chao ◽

Jingxuan Tian ◽

...

Keyword(s):

Phase Separation ◽

Aqueous Phase

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Phase separation at the nanoscale quantified by dcFCCS

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2008447117 ◽

2020 ◽

Vol 117 (44) ◽

pp. 27124-27131

Author(s):

Sijia Peng ◽

Weiping Li ◽

Yirong Yao ◽

Wenjing Xing ◽

Pilong Li ◽

...

Keyword(s):

Phase Separation ◽

Detection Limit ◽

Fluorescence Microscopy ◽

Critical Concentration ◽

Transition Process ◽

Correlation Spectroscopy ◽

Cellular Processes ◽

Conventional Fluorescence Microscopy ◽

Micrometer Scale ◽

Liquid Liquid Phase Separation

Liquid–liquid phase separation, driven by multivalent macromolecular interactions, causes formation of membraneless compartments, which are biomolecular condensates containing concentrated macromolecules. These condensates are essential in diverse cellular processes. Formation and dynamics of micrometer-scale phase-separated condensates are examined routinely. However, limited by commonly used methods which cannot capture small-sized free-diffusing condensates, the transition process from miscible individual molecules to micrometer-scale condensates is mostly unknown. Herein, with a dual-color fluorescence cross-correlation spectroscopy (dcFCCS) method, we captured formation of nanoscale condensates beyond the detection limit of conventional fluorescence microscopy. In addition, dcFCCS is able to quantify size and growth rate of condensates as well as molecular stoichiometry and binding affinity of client molecules within condensates. The critical concentration to form nanoscale condensates, identified by our experimental measurements and Monte Carlo simulations, is at least several fold lower than the detection limit of conventional fluorescence microscopy. Our results emphasize that, in addition to micrometer-scale condensates, nanoscale condensates are likely to play important roles in various cellular processes and dcFCCS is a simple and powerful quantitative tool to examine them in detail.

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