Hello, Your question was forwarded to chimera-users@cgl.ucsf.edu (the recommended address for questions about Chimera). (1) To build a structure of a fusion protein, first you would need to have the structures of the individual proteins. You can search the Protein DataBank (PDB) to see if they are available, e.g. directly using their website <https://www.rcsb.org/> ... or you can search the PDB sequences with your sequence using the Blast Protein tool (in Chimera menu under Tools... Sequence). <https://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/blast.html> If the exact same proteins that you need are not available as experimentally solved structures in the Protein DataBank, you may be able to get comparative models for them from ModBase, <https://modbase.compbio.ucsf.edu/modbase-cgi/index.cgi> ...or create your own comparative model in Chimera <https://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/multalignviewer/modeller.html#comparative> <https://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/tutorials/dor.html> (2) Open the structures (PDB files) in Chimera. They will be two different models, e.g. #0 and #1, in Chimera. (3) Delete any parts of the structures you don't want. One way: if you show sequence (Chimera menu: Favorites... Sequence...) of the model, you can drag a box in the sequence to select the part of the structure that you don't want, then use main menu Actions... Atoms/Bonds... delete. There is no undo for deleting, so if you mess up, you need to close the model and reopen the original PDB file. (4) Then if you already have all the residues needed for your fusion protein, you can join the C-term of a chain in one model to the N-term of a chain in the other model to create a new model, #2. To do this you would use the Build Structure tool (in menu under Tools... Structure Editing), the Join Models section. <https://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/editing/editing.html#join> To be able to select the two atoms to join, you need to hide ribbons and show the backbone atoms, e.g. commands: ~ribbon ~disp display @n,ca,c,o If the ribbon is shown, normally you can't see the backbone atoms. To select the two atoms from the screen, you would Ctrl-click one, Shift-Ctrl-click the other. <https://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/selection.html> (3b) If you needed to add some linker residues between, however, you would do that before joining. You could do it by adding amino acids to the end of one of the models with the "addaa" command, see: <https://www.rbvi.ucsf.edu/chimera/docs/UsersGuide/midas/addaa.html> ...or another way is to just build the linker segment as a separate peptide with Build Structure, start structure, peptide <https://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/editing/editing.html#start> ...then use two joining steps as in (4) above to put the three parts into one model. Currently we are no longer developing Chimera, and recommend using ChimeraX instead for most purposes. ChimeraX has all of the same features as mentioned above for Chimera, except not "addaa," but you could build a linker as a separate peptide as mentioned above. ChimeraX is generally easier to use, has nicer rendering, is faster on big structures, etc. <https://www.rbvi.ucsf.edu/chimerax/index.html> <https://www.rbvi.ucsf.edu/chimerax/docs/user/advantages.html> I hope this helps, Elaine ----- Elaine C. Meng, Ph.D. UCSF Chimera(X) team Department of Pharmaceutical Chemistry University of California, San Francisco

On Aug 4, 2022, at 7:40 AM, Eric Pettersen via Chimera-users <chimera-users@cgl.ucsf.edu> wrote:

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From: Milad Mapar <milad.mapar@yahoo.com> Subject: Making Fusion Proteins using Chimera Date: August 4, 2022 at 6:26:46 AM PDT

Hello With thanks to your support on Chimera, could you please help me to make a Fusion Protein using Chimera 1.16 Any links to tutorials or hints would be appreciated Many thanks

Pharmaceutical Biotechnology PhD Student Milad