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Rapid Enrichment of a Native Multipass Transmembrane Protein via Cell Membrane Electrophoresis through Buffer pH and Ionic Strength Adjustment.
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- Author(s): Liu TT;Liu TT; Huang SH; Huang SH; Chao L; Chao L
- Source:
Journal of the American Chemical Society [J Am Chem Soc] 2024 May 01; Vol. 146 (17), pp. 11634-11647. Date of Electronic Publication: 2024 Apr 17.
- Publication Type:
Journal Article; Research Support, Non-U.S. Gov't; Research Support, N.I.H., Extramural; Research Support, U.S. Gov't, Non-P.H.S.
- Language:
English
- Additional Information
- Source:
Publisher: American Chemical Society Country of Publication: United States NLM ID: 7503056 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1520-5126 (Electronic) Linking ISSN: 00027863 NLM ISO Abbreviation: J Am Chem Soc Subsets: MEDLINE
- Publication Information:
Publication: Washington, DC : American Chemical Society
Original Publication: Easton, Pa. [etc.]
- Subject Terms:
- Abstract:
Supported membrane electrophoresis is a promising technique for collecting membrane proteins in native bilayer environments. However, the slow mobility of typical transmembrane proteins has impeded the technique's advancement. Here, we successfully applied cell membrane electrophoresis to rapidly enrich a 12-transmembrane helix protein, glucose transporter 1 with antibodies (GLUT1 complex), by tuning the buffer pH and ionic strength. The identified conditions allowed the separation of the GLUT1 complex and a lipid probe, Fast-DiO, within a native-like environment in a few minutes. A force model was developed to account for distinct electric and drag forces acting on the transmembrane and aqueous-exposed portion of a transmembrane protein as well as the electroosmotic force. This model not only elucidates the impact of size and charge properties of transmembrane proteins but also highlights the influence of pH and ionic strength on the driving forces and, consequently, electrophoretic mobility. Model predictions align well with experimentally measured electrophoretic mobilities of the GLUT1 complex and Fast-DiO at various pH and ionic strengths as well as with several lipid probes, lipid-anchored proteins, and reconstituted membrane proteins from previous studies. Force analyses revealed the substantial membrane drag of the GLUT1 complex, significantly slowing down electrophoretic mobility. Besides, the counterbalance of similar magnitudes of electroosmotic and electric forces results in a small net driving force and, consequently, reduced mobility under typical neutral pH conditions. Our results further highlight how the size and charge properties of transmembrane proteins influence the suitable range of operating conditions for effective movement, providing potential applications for concentrating and isolating membrane proteins within this platform.
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- Accession Number:
0 (Membrane Proteins)
0 (Buffers)
0 (Glucose Transporter Type 1)
- Publication Date:
Date Created: 20240417 Date Completed: 20240501 Latest Revision: 20240505
- Publication Date:
20240505
- Accession Number:
PMC11066866
- Accession Number:
10.1021/jacs.3c13579
- Accession Number:
38628144
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