Furukawa, H, Simorowski, N, Michalski, K (January 2021) Effective production of oligomeric membrane proteins by EarlyBac-insect cell system. In: Methods in Enzymology. Elsevier.
Abstract
Despite major advances in methodologies for membrane protein production over the last two decades, there remain challenging protein complexes that are technically difficult to yield by conventional recombinant expression methods. A large number of these proteins are multimeric membrane proteins from eukaryotic species, which are required to pass through stringent quality control mechanisms of host cells for proper folding and complex assembly. Here, we describe the development procedure to improve the production efficiency of multi-oligomeric membrane protein complexes in insect cells and recombinant baculovirus, which involves screening of promoters, enhancers, and untranslated regions for expression levels, using calcium homeostasis modulator (CALHM) and N-methyl-D-aspartate receptor (NMDAR) proteins as examples. We demonstrate that our insect cell expression strategy is effective in expression of both multi-homomeric CALHM proteins and multi-heteromeric NMDARs.
| Item Type: | Book Section |
|---|---|
| Subjects: | Investigative techniques and equipment > microscopy > Cryo-electron microscopy bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > NMDA receptor organs, tissues, organelles, cell types and functions > sub-cellular tissues: types and functions > calcium channel Investigative techniques and equipment > EarlyBac Investigative techniques and equipment > x ray crystallography |
| CSHL Authors: | |
| Communities: | CSHL labs > Furukawa lab |
| SWORD Depositor: | CSHL Elements |
| Depositing User: | CSHL Elements |
| Date: | 1 January 2021 |
| Date Deposited: | 06 May 2021 13:48 |
| Last Modified: | 07 Mar 2022 14:32 |
| PMCID: | PMC8896536 |
| URI: | https://repository.cshl.edu/id/eprint/39998 |
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