Palindromic sequences involved in links.

[atroel01]

XiSearch analytical problem:

I am doing a small crosslinking for structure experiment and there is a palindromic sequence of KKLLKK which is involved in both DSSO and EDC type crosslinks. The default reverse decoy results in a somewhat concerning number of topmatches being the REVerse decoy of this sequence ie KKLLK, KLLK or LLKK- which are obviously exactly the same the same as forward in terms of sequence.

Does xiSearch deal with these edge cases in some way?

Unclear instructions or a Bug

When i provide a shuffled fasta instead, appending REV_ in front of the protein name, it ignores it and instead generates a a new REV_ and even a REV_REV_ anyway!

Attempted Mitigations:

Tested in both 1.8.7 and 1.8.11.
Submitted both as single FASTA and as two separate fastas Indicating the decoy on the shuffled FASTA.

Question

What is the appropriate way to deal with these kinds of sequences?

Thanks

ADDIT

After furhter searching through source code there is a custom Settings to addres this:

AUTODECOY:false
DECOY_GENERATION:shuffle

[lutzfischer]

AUTODECOY:false 

disables all automatic decoy generation. But probably the better way for you is to have your decoys in a separate FASTA file and tick the decoy checkbox on the decoy FASTA. This way xiSEARCH is aware that you made your own decoys and also does not autogenerate any. One advantage over just adding AUTODECOY:false is that xiSEARCH still flags decoy matches as such.

Also you have several options for randomized FASTA instead of reversed. will mix up the sequence of each protein.

Options you have:

Mix up the sequence of each protein with only the aminoacids of that protein

DECOY_GENERATION:shuffle

Completely randomly generate proteins of the same length:

DECOY_GENERATION:random

Randomly generate the decoy proteins but the possible aminoacids are taken for each target protein and the pair wise propability of amino acids following each other is taken into account

DECOY_GENERATION:random_directed

However by default the decoy generation is digestion aware. And for all but the reversed methods this means that the digestible amino acids stay in place - meaning All Ks and Rs stay where they are, which is probably still ok as it would be unlikely to still have the LL in place. You can switch that of with

DECOY_DIGESTION_AWARE:false

[lutzfischer]

If you want to use one or more of these option regularly you could add them to the BasicConfigEntries.conf file in the xiSEARCH folder.
The last section, [CUSTOM], has options that can be selected when pressing the [+] button besides the custom settings

you could add things like:

Don't automaticaly genereate decoys=AUTODECOY:false

If you like to have comments in your config describing what it does you can add this there as well:

Don't automaticaly genereate decoys # Do not automatically generate decoys - be aware that you should supply your own decoys#=AUTODECOY:false

Would add:

# Do not automatically generate decoys - be aware that you should supply your own decoys
AUTODECOY:false

to the config.

[atroel01]

Thanks!

Even though I did supply two fastas with one being decoy I still encountered the problem.
I had wondered if it was due to parsing of headers for decoys- but I used the stand alone tool to parse my fasta and make a shuffled decoy, supplying both with decoy ticked. It Still it continued to make decoys of my decoys.

It would be good to have some of these custom commands documented or at least by default included in the configs.

[lutzfischer]

Ok having ticked decoy and it still does generate decoys sounds like a bug - will have a look.