DirectMX - One-Step Reconstitution of Membrane Proteins From Crude Cell Membranes Into Salipro Nanoparticles
Integral membrane proteins (IMPs) are central to many physiological processes and represent ∼60% of current drug targets. An intricate interplay with the lipid molecules in the cell membrane is known to influence the stability, structure and function of IMPs. Detergents are commonly used to solubili...
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Published in | Frontiers in bioengineering and biotechnology Vol. 8; p. 215 |
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Main Authors | , , , , , |
Format | Journal Article |
Language | English |
Published |
Switzerland
Frontiers Media S.A
19.03.2020
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Subjects | |
Online Access | Get full text |
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Summary: | Integral membrane proteins (IMPs) are central to many physiological processes and represent ∼60% of current drug targets. An intricate interplay with the lipid molecules in the cell membrane is known to influence the stability, structure and function of IMPs. Detergents are commonly used to solubilize and extract IMPs from cell membranes. However, due to the loss of the lipid environment, IMPs usually tend to be unstable and lose function in the continuous presence of detergent. To overcome this problem, various technologies have been developed, including protein engineering by mutagenesis to improve IMP stability, as well as methods to reconstitute IMPs into detergent-free entities, such as nanodiscs based on apolipoprotein A or its membrane scaffold protein (MSP) derivatives, amphipols, and styrene-maleic acid copolymer-lipid particles (SMALPs). Although significant progress has been made in this field, working with inherently unstable human IMP targets (e.g., GPCRs, ion channels and transporters) remains a challenging task. Here, we present a novel methodology, termed DirectMX (for direct membrane extraction), taking advantage of the saposin-lipoprotein (Salipro) nanoparticle technology to reconstitute fragile IMPs directly from human crude cell membranes. We demonstrate the applicability of the DirectMX methodology by the reconstitution of a human solute carrier transporter and a wild-type GPCR belonging to the human chemokine receptor (CKR) family. We envision that DirectMX bears the potential to enable studies of IMPs that so far remained inaccessible to other solubilization, stabilization or reconstitution methods. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 These authors have contributed equally to this work This article was submitted to Synthetic Biology, a section of the journal Frontiers in Bioengineering and Biotechnology Reviewed by: Albert Guskov, University of Groningen, Netherlands; Cédric Orelle, UMR5086 Microbiologie Moléculaire et Biochimie Structurale (MMSB), France Edited by: Alexej Kedrov, Heinrich Heine University Düsseldorf, Germany |
ISSN: | 2296-4185 2296-4185 |
DOI: | 10.3389/fbioe.2020.00215 |