Capillary Zone Electrophoresis-Tandem Mass Spectrometry for Top-Down Proteomics of Mouse Brain Integral Membrane Proteins.
Anal Chem
; 95(34): 12590-12594, 2023 08 29.
Article
in En
| MEDLINE
| ID: mdl-37595263
ABSTRACT
Mass spectrometry (MS)-based top-down characterization of integral membrane proteins (IMPs) is crucial for understanding their functions in biological processes. However, it is technically challenging due to their low solubility in typical MS-compatible buffers. In this work, for the first time, we developed an efficient capillary zone electrophoresis (CZE)-tandem MS (MS/MS) method for the top-down proteomics (TDP) of IMPs enriched from mouse brains. Our technique employs a sample buffer containing 30% (v/v) formic acid and 60% (v/v) methanol for solubilizing IMPs and utilizes a separation buffer of 30% (v/v) acetic acid and 30% (v/v) methanol for maintaining the solubility of IMPs during CZE separation. Single-shot CZE-MS/MS identified 51 IMP proteoforms from the mouse brain sample. Coupling size exclusion chromatography (SEC) to CZE-MS/MS enabled the identification of 276 IMP proteoforms from the mouse brain sample containing 1-4 transmembrane domains. This proof-of-concept work demonstrates the high potential of CZE-MS/MS for the large-scale TDP of IMPs.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Methanol
/
Tandem Mass Spectrometry
Limits:
Animals
Language:
En
Journal:
Anal Chem
Year:
2023
Type:
Article
Affiliation country:
United States