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  Top-Down Identification and Sequence Analysis of Small Membrane Proteins Using MALDI-MS/MS

Meier-Credo, J., Preiss, L., Wüllenweber, I., Resemann, A., Nordmann, C., Zabret, J., et al. (2022). Top-Down Identification and Sequence Analysis of Small Membrane Proteins Using MALDI-MS/MS. Journal of the American Society for Mass Spectrometry, 33(7), 1293-1302. doi:10.1021/jasms.2c00102.

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Meier-Credo, Jakob1, 2, Author           
Preiss, Laura3, Author           
Wüllenweber, Imke1, 2, Author           
Resemann, Anja4, Author
Nordmann, Christoph4, Author
Zabret, Jure5, Author
Suckau, Detlef4, Author
Michel, Hartmut6, Author                 
Nowacyk, Marc M.5, Author
Meier, Thomas7, Author
Langer, Julian David1, 2, Author                 
1Proteomics and Mass Spectrometry, Max Planck Institute of Biophysics, Max Planck Society, ou_3262216              
2Proteomics, Max Planck Institute for Brain Research, 60438 Frankfurt am Main, Germany, ou_persistent22              
3Department of Structural Biology, Max Planck Institute of Biophysics, Max Planck Society, ou_2068291              
4Bruker Daltonics GmbH & Co. KG, Bremen, Germany, Fahrenheitstrasse 4, 28359 Bremen, Germany, ou_persistent22              
5Department of Plant Biochemistry, Ruhr University Bochum, 44780 Bochum, Germany, ou_persistent22              
6Department of Molecular Membrane Biology, Max Planck Institute of Biophysics, Max Planck Society, ou_2068290              
7Department of Life Sciences, Imperial College London, Exhibition Road, SW7 2AZ London, United Kingdom, ou_persistent22              


Free keywords: Extraction,Genetics, Mass spectrometry, Peptides and proteins, Protein identification
 Abstract: Identification and sequence determination by mass spectrometry have become routine analyses for soluble proteins. Membrane proteins, however, remain challenging targets due to their hydrophobicity and poor annotation. In particular small membrane proteins often remain unnoticed as they are largely inaccessible to Bottom-Up proteomics. Recent advances in structural biology, though, have led to multiple membrane protein complex structures being determined at sufficiently high resolution to detect uncharacterized, small subunits. In this work we offer a guide for the mass spectrometric characterization of solvent extraction-based purifications of small membrane proteins isolated from protein complexes and cellular membranes. We first demonstrate our Top-Down MALDI-MS/MS approach on a Photosystem II preparation, analyzing target protein masses between 2.5 and 9 kDa with high accuracy and sensitivity. Then we apply our technique to purify and sequence the mycobacterial ATP synthase c subunit, the molecular target of the antibiotic drug bedaquiline. We show that our approach can be used to directly track and pinpoint single amino acid mutations that lead to antibiotic resistance in only 4 h. While not applicable as a high-throughput pipeline, our MALDI-MS/MS and ISD-based approach can identify and provide valuable sequence information on small membrane proteins, which are inaccessible to conventional Bottom-Up techniques. We show that our approach can be used to unambiguously identify single-point mutations leading to antibiotic resistance in mycobacteria.


Language(s): eng - English
 Dates: 2022-06-152022-04-172022-06-152022-06-272022-07-06
 Publication Status: Published in print
 Pages: 10
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1021/jasms.2c00102
PMID: 35758524
 Degree: -



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Source 1

Title: Journal of the American Society for Mass Spectrometry
Source Genre: Journal
Publ. Info: New York, NY : Springer
Pages: - Volume / Issue: 33 (7) Sequence Number: - Start / End Page: 1293 - 1302 Identifier: ISSN: 1044-0305
CoNE: https://pure.mpg.de/cone/journals/resource/954925590403