MolecularCloud January Newsletter: Detergent-free Extraction of Membrane Protein
The preparation of samples exhibiting stable and natively-folded structures that retain the function of the membrane proteins is crucial for investigation of high-resolution structures and dynamics of membrane proteins.
Detergents and micelles, as traditional membrane mimetics, are continued to be used in the structural studies, mainly the X-ray crystallography studies. However, it is known that detergents destabilize protein structures and disrupt their functions. Bicelles, although considered to be better membrane mimetic systems, have also been shown to distort protein structures and functions due to the diffusion of the detergent molecules from the bicelle rim to the planar lipid bilayer region.
Dr. Bankala Krishnarjuna’s team published an article titled “Detergent-free extraction, reconstitution and characterization of membrane-anchored cytochrome-b5 in native lipids” in Chemical Communications, introducing a novel method for membrane protein extraction without destabilizing protein structures (Chemical Communications, 2020). In this study, the researchers demonstrated the feasibility of a direct extraction of a B16 kDa cytochrome-b5 from E. coli using a synthetic styrene-co-maleic acid-ethanolamine (SMA-EA) polymer and reconstituting the cytochrome-b5 protein in polymer nanodiscs along with native lipids from E. coli.
A schematic representation of the direct extraction of a 16 kDa rabbit cytochrome-b5 in native E. coli lipids using a negatively charged SMA-EA polymer. + and - indicate the net charge of a membrane protein. SEC: size-exclusion chromatography.
The result data of this research revealed that the directly extracted 15N-labelled cytochrome-b5 was of high purity as indicated by the 2D 1H/15N TROSY-HSQC NMR spectrum, which also demonstrated the correctly folded state of the protein and the feasibility to carry out structural and dynamical measurements using the well-established multidimensional solution NMR techniques.
Time-limited Campaign
Is extraction using synthetic polymers a best way for preparing membrane protein samples? Leave your thoughts in the comment section below to win a canvas bag, hoodie or $20 Amazon gift card! You can also have a chance to get one of those gifts by taking the short quiz below from Dr. Bankala regarding membrane protein extraction.
*MolecularCloud reserves the right of final explanation for details of this campaign. If there's any further question, please feel free to contact us at support@molecularcloud.org.
Previous Newsletters
MolecularCloud December Newsletter: Methods and Protocols for Cereal Genomics
MolecularCloud November Newsletter: Predicting the effect of mutations on protein–RNA bindingMolecularCloud October Newsletter: Novel Base Editors Induce Efficient and Specific Editing with Low DDR
Q1: How many membrane protein structures have been reported in the Protein Data Bank? (1 mark)
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15% of all reported protein structures11
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10% of all reported protein structures13
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5% of all reported protein structures9
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<3% of all reported protein structures30
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Q2: Is it possible to extract the native lipids associated with a specific membrane protein in the cell membrane? (2 marks)
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Yes48
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No10
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Q3: Which of the following can achieve 100% detergent-free extraction of membrane proteins? (3 marks)
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Detergents2
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Micelles8
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Synthetic polymers48
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Q4: Are membrane proteins extracted using synthetic polymers suitable for high-resolution structural studies by NMR (4 marks)?
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Yes47
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No11
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Q5: Is the charge of the polymer used a limitation for achieving membrane protein extraction and reconstitution? (5 marks)
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Yes34
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No24
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Can polymers be used also to isolate proteins from non-membrane organelles?
Spectacular data. Can't wait to see this method to be reproduced by other groups.
I would not call it as a new method. However, I am excited to see the refinement of the method! Well done!
GOOD METHOD
On my way. I’ll try it, but how to get the polymers.
GOOD METHOD
I would not call it as a new method. However, I am excited to see the refinement of the method! Well done!
Could be useful also for our research.
Sounds useful.
Exciting field!