Glycosyl Phosphatidylinositol-Linked Membrane Proteins: Structure, Biosynthesis, and Function

AbstractCellular membranes and associated proteins play critical physiological roles in organisms from all life kingdoms. In many cases, malfunction of biological membranes triggered by changes in the lipid bilayer properties or membrane protein functional abnormalities lead to severe diseases. To understand in detail the processes that govern the life of cells and to control diseases, one of the major tasks in biological sciences is to learn how the membrane proteins function. To do so, a variety of biochemical and biophysical approaches have been used in molecular studies of membrane protein structure and function on the nanoscale. This review focuses on electron paramagnetic resonance with site-directed nitroxide spin-labeling (SDSL EPR), which is a rapidly expanding and powerful technique reporting on the local protein/spin-label dynamics and on large functionally important structural rearrangements. On the other hand, adequate to nanoscale study membrane mimetics have been developed and used in conjunction with SDSL EPR. Primarily, these mimetics include various liposomes, bicelles, and nanodiscs. This review provides a basic description of the EPR methods, continuous-wave and pulse, applied to spin-labeled proteins, and highlights several representative applications of EPR to liposome-, bicelle-, or nanodisc-reconstituted membrane proteins.

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Influence of Detergent Properties on the Solubilization and Function of Membrane Proteins

Biophysical Journal ◽

10.1016/j.bpj.2013.11.1298 ◽

2014 ◽

Vol 106 (2) ◽

pp. 222a

Author(s):

Ashton T. Brock ◽

Linda Columbus

Keyword(s):

Membrane Proteins ◽

And Function

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Glycosyl-phosphatidylinositol-anchored membrane proteins can be distinguished from transmembrane polypeptide-anchored proteins by differential solubilization and temperature-induced phase separation in Triton X-114

Biochemical Journal ◽

10.1042/bj2800745 ◽

1991 ◽

Vol 280 (3) ◽

pp. 745-751 ◽

Cited By ~ 55

Author(s):

N M Hooper ◽

A Bashir

Keyword(s):

Phase Separation ◽

Membrane Proteins ◽

Aqueous Phase ◽

Hydrophobic Membrane ◽

Rich Phase ◽

Glycosyl Phosphatidylinositol ◽

Ionic Detergent ◽

Membrane Spanning ◽

Triton X ◽

Insoluble Pellet

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 Relationship between Structural Symmetry and Function in Membrane Proteins

Biophysical Journal ◽

10.1016/j.bpj.2020.11.410 ◽

2021 ◽

Vol 120 (3) ◽

pp. 24a-25a

Author(s):

Emily L. Yaklich ◽

Antoniya A. Aleksandrova ◽

Lucy R. Forrest

Keyword(s):

Membrane Proteins ◽

Structural Symmetry ◽

And Function ◽

The Relationship

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Correlated STORM-homoFRET Imaging: Applications to the Study of Membrane Receptor Self-Association and Clustering

10.1101/2021.12.21.473581 ◽

2021 ◽

Author(s):

Amine Driouchi ◽

Scott Gray-Owen ◽

Christopher M Yip

Keyword(s):

Spatial Distribution ◽

Membrane Proteins ◽

Complex Mixture ◽

Cell Surface Receptor ◽

Oligomeric State ◽

Imaging Approach ◽

Surface Receptor ◽

A Cell ◽

Self Association ◽

And Function

Mapping the self-organization and spatial distribution of membrane proteins is key to understanding their function. We report here on a correlated STORM/homoFRET imaging approach for resolving the nanoscale distribution and oligomeric state of membrane proteins. Live cell homoFRET imaging of CEACAM1, a cell-surface receptor known to exist in a complex equilibrium between monomer and dimer/oligomer states, revealed highly heterogenous diffraction-limited structures on the surface of HeLa cells. Correlated super-resolved STORM imaging revealed that these structures comprised a complex mixture and spatial distribution of self-associated CEACAM1 molecules. This correlated approach provides a compelling strategy for addressing challenging questions about the interplay between membrane protein concentration, distribution, interaction, clustering, and function.

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Bioinformatic approaches for the structure and function of membrane proteins

BMB Reports ◽

10.5483/bmbrep.2009.42.11.697 ◽

2009 ◽

Vol 42 (11) ◽

pp. 697-704 ◽

Cited By ~ 17

Author(s):

Hyun-Jun Nam ◽

Jou-Hyun Jeon ◽

Sang-Uk Kim

Keyword(s):

Membrane Proteins ◽

Structure And Function ◽

Bioinformatic Approaches ◽

And Function

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Structure and function of qiuinone binding membrane proteins

Membrane Proteins - Advances in Protein Chemistry ◽

10.1016/s0065-3233(03)63007-x ◽

2003 ◽

pp. 151-176 ◽

Cited By ~ 3

Author(s):

Momi Iwata ◽

Jeff Abramson ◽

Bernadette Byrne ◽

S.O Iwata

Keyword(s):

Membrane Proteins ◽

Structure And Function ◽

And Function

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Biophysical methods toolbox to study ABC exporter structure and function

Biological Chemistry ◽

10.1515/hsz-2016-0244 ◽

2017 ◽

Vol 398 (2) ◽

pp. 229-235

Author(s):

Thomas Marcellino ◽

Vasundara Srinivasan

Keyword(s):

Membrane Proteins ◽

Abc Transporter ◽

Integrated Approach ◽

Structure And Function ◽

Dynamic Membrane ◽

Biophysical Characterization ◽

Intermediate States ◽

And Function

Abstract ABC exporters are highly dynamic membrane proteins that span a huge spectrum of different conformations. A detailed integrated approach of cellular, biochemical and biophysical characterization of these ‘open’, ‘closed’ and other intermediate states is central to understanding their function. Almost 40 years after the discovery of the first ABC transporter, thanks to the enormous development in methodologies, a picture is slowly emerging to visualize how these fascinating molecules transport their substrates. This mini review summarizes some of the biophysical tools that have made a major impact in understanding the function of the ABC exporters.

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