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| Item Type: | Article |
|---|---|
| Title: | Delineation of functional subdomains of Huntingtin protein and their interaction with HAP40 |
| Creators Name: | Alteen, M.G., Deme, J.C., Alvarez, C.P., Loppnau, P., Hutchinson, A., Seitova, A., Chandrasekaran, R., Silva Ramos, E., Secker, C., Alqazzaz, M., Wanker, E.E., Lea, S.M., Arrowsmith, C.H. and Harding, R.J. |
| Abstract: | The huntingtin (HTT) protein plays critical roles in numerous cellular pathways by functioning as a scaffold for its many interaction partners and HTT knock out is embryonic lethal. Interrogation of HTT function is complicated by the large size of this protein so we studied a suite of structure-rationalized subdomains to investigate the structure-function relationships within the HTT-HAP40 complex. Protein samples derived from the subdomain constructs were validated using biophysical methods and cryo-electron microscopy, revealing they are natively folded and can complex with validated binding partner, HAP40. Derivatized versions of these constructs enable protein-protein interaction assays in vitro, with biotin tags, and in cells, with luciferase two-hybrid assay-based tags, which we use in proof-of-principle analyses to further interrogate the HTT-HAP40 interaction. These open-source biochemical tools enable studies of fundamental HTT biochemistry and biology, will aid the discovery of macromolecular or small-molecule binding partners and help map interaction sites across this large protein. |
| Keywords: | Huntington’s Disease, Neurodegeneration, Protein Purification, Protein Structure, Cryo-Electron Microscopy, Protein-Protein Interaction |
| Source: | Structure |
| ISSN: | 0969-2126 |
| Publisher: | Cell Press |
| Volume: | 31 |
| Number: | 9 |
| Page Range: | 1121-1131.e6 |
| Date: | 7 September 2023 |
| Additional Information: | Copyright © 2023 Elsevier Ltd. All rights reserved. |
| Official Publication: | https://doi.org/10.1016/j.str.2023.06.002 |
| External Fulltext: | View full text on external repository or document server |
| PubMed: | View item in PubMed |
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