Multicolour fluorescence-detection size-exclusion chromatography for structural genomics of membrane multiprotein complexes
- Many interesting and important membrane proteins are hetero-oligomeric. However, besides naturally abundant examples, the structures of relatively few such complexes are known. Partly, this is due to difficulties in expression, stoichiometric assembly, and in the evaluation of their stability prior to crystallization trials. Here we describe a new approach, which allows rapid assessment of protein complex quality, assembly and stoichiometry, simplifying the search for conditions conducive to long-term stability and crystallization. Multicolour fluorescence size-exclusion chromatography (MC-FSEC) is used to enable tracking of individual subunits through expression, solubilization and purification steps. We show how the method has been applied to the heterodimeric transporter associated with antigen processing (TAP) and demonstrate how it may be extended in order to analyse membrane multisubunit assemblies.
| Author: | David Parcej, Renate Guntrum, Sabine Schmidt, Andreas Hinz, Robert TampéORCiDGND |
|---|---|
| URN: | urn:nbn:de:hebis:30:3-305907 |
| DOI: | https://doi.org/10.1371/journal.pone.0067112 |
| ISSN: | 1932-6203 |
| Pubmed Id: | https://pubmed.ncbi.nlm.nih.gov/23825631 |
| Parent Title (English): | PLoS One |
| Publisher: | PLoS |
| Place of publication: | Lawrence, Kan. |
| Document Type: | Article |
| Language: | English |
| Date of Publication (online): | 2013/06/25 |
| Date of first Publication: | 2013/06/25 |
| Publishing Institution: | Universitätsbibliothek Johann Christian Senckenberg |
| Release Date: | 2013/07/01 |
| Volume: | 8 |
| Issue: | (6):e67112 |
| Page Number: | 9 |
| Note: | Copyright: © 2013 Parcej et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
| HeBIS-PPN: | 347477712 |
| Institutes: | Biochemie, Chemie und Pharmazie / Biochemie und Chemie |
| Dewey Decimal Classification: | 5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie |
| Sammlungen: | Universitätspublikationen |
| Sammlung Biologie / Sondersammelgebiets-Volltexte | |
| Licence (German): |  Creative Commons - Namensnennung 3.0 |
