Hello everyone,

I'm very new to using Rosetta and I'm having problems generating 3mers and 9mers. I've followed the tutorials and the documentation, but I continue to receive an error.

I've created a directory (input_files) with the following files: a FASTA sequence, an ss2 file (generated using PsiPred), and a PDB file (I've tried using both the one generated by AlphaFold3 and the one generated by DMPFold prediction but it doesn't seem to make a difference). The protein has 322 amino acids and I've verified that all three input files have 322 amino acids.

This is the command I use in the shell:

../../rosetta.binary.ubuntu.release-371/main/source/build/src/release/linux/5.4/64/x86/gcc/7/static/fragment_picker.static.linuxgccrelease @flags

And this is the contenute of "flags":

# Input databases
-in::file::vall                 ../../rosetta.binary.ubuntu.release-371/main/tools/fragment_tools/vall.apr24.2008.extended.gz

# Query-related input files
#-in::file::checkpoint          input_files/0296.checkpoint
-in::file::fasta                input_files/0296.fasta
-in::file::s                    input_files/0296.pdb
-frags::ss_pred                 input_files/0296.psipred.ss2 predA

# Weights file
-frags::scoring::config         input_files/simple.wghts

# What should we do?
-frags::bounded_protocol

# three-mers and nine-mers
-frags::frag_sizes              3 9
-frags::n_candidates            200
-frags::n_frags                 200

# Output
-out::file::frag_prefix         output_files/frags
-frags::describe_fragments      output_files/frags.fsc

This is the simple.wghts file:

# score name          priority  wght   max_allowed  extras
RamaScore               400    2.0     -       predA
SecondarySimilarity     350     1.0            predA
FragmentCrmsd             0     0.0     -

and this is the error in crash report: 

[FILE]: src/protocols/frag_picker/scores/SecondarySimilarity.hh
[LINE]: 99
[START_MESSAGE]
[ ERROR ] UtilityExitException
ERROR: Unable to find secondary structure prediction for psipred

Lines 97-99 of SecondarySimilarity.hh are:

core::fragment::SecondaryStructureOP query_prediction( picker->get_query_ss(prediction_id) );
                if ( ! query_prediction ) {
                        utility_exit_with_message("Unable to find secondary structure prediction for " + prediction_id );

If I use, instead of simple.wghts:

SecondarySimilarity      350     0.5 - predA
ProfileScoreL1           50      1 -
FragmentCrmsd            30      0.0 -
RamaScore                100     2.0 - predA

this is the error:

[FILE]: src/protocols/frag_picker/scores/SecondarySimilarity.cc
[LINE]: 136
[START_MESSAGE]
[ ERROR ] UtilityExitException
ERROR: Assertion `query_prediction->total_residue() == query_len_` failed.

Can you please help me making this fragment picker work?