starting sequence with flexible backbone design

[Anonymous]

Hi,

We have a question regarding the starting sequence during the sequence design process when you have an initial backbone structure but no initial sequence.

We have tried submitting a random sequence to the Robetta fragment server but this results in an error due the server being unable to find any fragments.

Now given that we already have a backbone structure and would only like Rosetta to make small adjustments to the backbone – can we safely just submit any existing protein sequence to Robetta to get fragments, edit the input PDB starting structure to have a random sequence, and assume that the fragments will be ignored in the design process?

Alternatively is there a better way to ensure we start sequence design from a random starting sequence?

thank you very much for you help in advance,
Kasia.

[bpierce]

Interesting. I wonder what your random sequence was that Robetta couldn’t find anything.
Anyway, scientifically speaking, here is what I would do. Make a sequence that is weighted toward the secondary structures of your starting backbone structure. eg use a lot of alanines for helical regions, hihg-beta propensity residues for your sheets, etc. This will be more work than making a random sequence, but it will give you fragments that are weighted towards the same secondary structure as your starting backbone, thereby biasing you to staying near that structure – making smaller adjustments as you want, rather than large random changes.
Also, you can make fragments yourself using the make_fragments script included with rosetta++. If you are up for doing that, you can play around with the sequences you use as input, and get different fragment files out – some may give you better design results than others. Just depends how much work you want to do.

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