Functional integrity of membrane protein rhodopsin solubilized by styrene-maleic acid copolymer

Pitch, Stephanie G.; Yao, Weekie; Szundi, István; Fay, Jonathan F.; Chen, Eefei; Shumate, Anthony; Kliger, David S.; Farrens, David

doi:10.1016/j.bpj.2021.05.008

Cited by 8 publications

(6 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Following extraction with SMA, MPs can later be reconstituted back into lipid bilayers for functional studies [ 119 , 122 ]. Proteins embedded in SMALPs can be further studied using a range of techniques, including mass spectrometry [ 135 , 137 , 138 , 141 ] and spectroscopy [ 115 , 116 , 122 , 126 , 128 , 139 , 142 ], and various structural methods [ 143 ], described below in detail.…”

Section: Applications Of Lipodiscs In Structural Biologymentioning

confidence: 99%

Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review

et al. 2022

View full text Add to dashboard Cite

Amphiphilic copolymers consisting of alternating hydrophilic and hydrophobic units account for a major recent methodical breakthrough in the investigations of membrane proteins. Styrene–maleic acid (SMA), diisobutylene–maleic acid (DIBMA), and related copolymers have been shown to extract membrane proteins directly from lipid membranes without the need for classical detergents. Within the particular experimental setup, they form disc-shaped nanoparticles with a narrow size distribution, which serve as a suitable platform for diverse kinds of spectroscopy and other biophysical techniques that require relatively small, homogeneous, water-soluble particles of separate membrane proteins in their native lipid environment. In recent years, copolymer-encased nanolipoparticles have been proven as suitable protein carriers for various structural biology applications, including cryo-electron microscopy (cryo-EM), small-angle scattering, and conventional and single-molecule X-ray diffraction experiments. Here, we review the current understanding of how such nanolipoparticles are formed and organized at the molecular level with an emphasis on their chemical diversity and factors affecting their size and solubilization efficiency.

show abstract

Section: Applications Of Lipodiscs In Structural Biologymentioning

confidence: 99%

Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review

et al. 2022

View full text Add to dashboard Cite

show abstract

“…It is possible that the local lipid content in the ‘annular shell’ for DIBMA–TSch would be enriched in these preparations compared to microsomes. Other polymer lipoprotein particle encapsulation studies observed impaired activation [ 23 ], and these results further suggest that the local lipid microenvironment could alter channel activation.…”

Section: Resultsmentioning

confidence: 66%

“…Interestingly, the state probability (Po) for the PKA-treated CFTR isolated under these conditions w (Figure 2b) at 37 °C, which is lower than the typically observed nearly locked ope with a Po of 0.96 for TSch in BHK microsomes fused to the same lipid bilayer [ possible that the local lipid content in the 'annular shell' for DIBMA-TSch would riched in these preparations compared to microsomes. Other polymer lipoprotein p encapsulation studies observed impaired activation [23], and these results further s that the local lipid microenvironment could alter channel activation.…”

Section: Dibma-tsch Assembly and Cryo-em Analysismentioning

confidence: 70%

Conformational Variability in Ground-State CFTR Lipoprotein Particle Cryo-EM Ensembles

Aleksandrov

Jensen

et al. 2022

IJMS

Self Cite

View full text Add to dashboard Cite

Cystic fibrosis transmembrane regulator (CFTR) is a dynamic membrane protein belonging to the ABC transporter family. It is unusual within this family as it is an ion channel, as opposed to a transporter. Activation of CFTR requires ATP and phosphorylation by PKA, and dysregulation of CFTR mediated salt and water homeostasis can lead to cystic fibrosis. Recent advancements in structural biological methods have led to more than 10 published CFTR structures, and, so far, all of these structures of CFTR, determined by cryo-EM, have been limited to detergent-purified protein preparations. To visualize CFTR in an environment that more closely represents its native membranous environment, we utilized two different lipoprotein particle encapsulation techniques: one in which the ion channel is first purified and then reconstituted using the membrane scaffolding protein Saposin A and another that uses the solubilizing polymer Sokalan CP9 (DIBMA) to extract CFTR directly from membranes. Structures derived from these types of preparations may better correlate to their function, for instance, the single-channel measurements from membrane vesicles.

show abstract

“…Many studies have reported that nanodiscs can be formed by exposing cell membranes or liposomes to SMA. [25][26][27][29][30][31][32][74][75][76] In the case of EVs, it is possible that increased membrane curvature combined with the glycocalyx makes them less susceptible to SMAinduced nanodisc formation than cell membranes or liposomes. Additionally, differences in the lipid composition and packaging between EVs and cells may also impact SMA membrane accessibility.…”

Section: Discussionmentioning

confidence: 99%

“…[25][26][27] SMA is primarily used to extract and study membrane proteins in their natural confirmation through solubilization as nanodiscs. [29][30][31][32] Although the copolymer has a broad solubilization capability, it exhibits varying degrees of effectiveness depending on membrane properties. [27,[33][34][35] For example, phospholipid bilayers containing more proteins pose a greater challenge for solubilization.…”

Section: Introductionmentioning

confidence: 99%

Chemically‐Induced Lipoprotein Breakdown for Improved Extracellular Vesicle Purification

Iannotta,

A.,

Lai

et al. 2023

Small

View full text Add to dashboard Cite

Extracellular vesicles (EVs) are nanosized biomolecular packages involved in intercellular communication. EVs are released by all cells, making them broadly applicable as therapeutic, diagnostic, and mechanistic components in (patho)physiology. Sample purity is critical for correctly attributing observed effects to EVs and for maximizing therapeutic and diagnostic performance. Lipoprotein contaminants represent a major challenge for sample purity. Lipoproteins are approximately six orders of magnitude more abundant in the blood circulation and overlap in size, shape, and density with EVs. This study represents the first example of an EV purification method based on the chemically‐induced breakdown of lipoproteins. Specifically, a styrene‐maleic acid (SMA) copolymer is used to selectively breakdown lipoproteins, enabling subsequent size‐based separation of the breakdown products from plasma EVs. The use of the polymer followed by tangential flow filtration or size‐exclusion chromatography results in improved EV yield, preservation of EV morphology, increased EV markers, and reduced contaminant markers. SMA‐based EV purification enables improved fluorescent labeling, reduces interactions with macrophages, and enhances accuracy, sensitivity, and specificity to detect EV biomarkers, indicating benefits for various downstream applications. In conclusion, SMA is a simple and effective method to improve the purity and yield of plasma‐derived EVs, which favorably impacts downstream applications.

show abstract

Functional integrity of membrane protein rhodopsin solubilized by styrene-maleic acid copolymer

Cited by 8 publications

References 42 publications

Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review

Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review

Conformational Variability in Ground-State CFTR Lipoprotein Particle Cryo-EM Ensembles

Chemically‐Induced Lipoprotein Breakdown for Improved Extracellular Vesicle Purification

Contact Info

Product

Resources

About