| Title |
EpicTope: predicting and validating non-disruptive epitope tagging sites |
| Authors |
Zinski, Joseph ; Joshi, Parnal ; Chung, Henri ; Preston, James ; Geller, Ari ; Warrick, Finn ; Berg, Brian D ; Tobin, Keagan ; Glova, Greg ; Liu, Fang ; Ming, Zhitao ; McGrail, Maura ; Balčiūnas, Darius ; Friedberg, Iddo ; Mullins, Mary C |
| DOI |
10.1242/dev.204502 |
| Full Text |
|
| Is Part of |
Development.. Cambridge : The Company of Biologists. 2026, vol. 153, iss. 5, p. 1-15.. ISSN 0950-1991. eISSN 1477-9129 |
| Keywords [eng] |
epitope tagging ; computational tool ; functional insertion sites ; zebrafish ; Smad5 ; Hdac1 ; protein engineering |
| Abstract [eng] |
Epitope tagging is a valuable technique enabling the in vivo identification, tracking and purification of proteins. We developed a tool, EpicTope, to facilitate this method by identifying amino acid positions most suitable for epitope insertion. Our method uses a scoring function that considers protein sequence secondary and tertiary structural features, solvent accessibility, and disordered binding regions to determine locations least disruptive to the protein function. We validated our approach on the zebrafish Smad5 and Hdac1 proteins. We show that multiple predicted internally tagged Smad5 proteins rescue zebrafish smad5 mutant embryos, while the N- and C-terminal-tagged variants do not, as predicted. Similarly, we found that optimally predicted internal and C-terminal Hdac1 tags rescued hdac1 mutant embryos, while a less-optimal N-terminal tag did not. We further show that these functionally tagged Smad5 and Hdac1 proteins are accessible to antibodies in whole-mount zebrafish embryo immunofluorescence, by western blotting and by immunoprecipitation from embryo extracts. Our work demonstrates that EpicTope is an accessible and effective tool for designing epitope tag insertion sites. |
| Published |
Cambridge : The Company of Biologists |
| Type |
Journal article |
| Language |
English |
| Publication date |
2026 |
| CC license |
|
