cryptogenomicon
commented
6 months ago Those two alignments don't appear to contain the same sequence data, so I wouldn't expect them to give the same profile. They aren't just the same alignment in two different formats.
Where you say "Input a3m created by removing all columns where the query had a gap and also removing deletions (lower-case letters) in non-query sequences so that everything has exactly the same length", you mean you're deleting residues from the (Stockholm version of the) alignment? That's changing the sequences; the two alignments are different after those steps are taken.
I've only glanced at this quickly, so if I've misunderstood something, let me know.
Augustin-Zidek
Hmmbuild gives different results depending on the input format. For Stockholm, it gives the output of the same length as the aligned sequences. For A3M, it sometimes does not.
In order to get the correctly aligned result, we therefore have to convert a3m -> sto - adding the
#=GC RFline while doing so - then run Hmmbuild with the converted Stockholm file.However, in our convertor, the
#=GC RFline is generated trivially:xin every position the first sequence of the a3m (i.e. the query sequence in the previous MSA search step) has an amino acid.in every column where the first sequence of the a3m has a gap (-).I realise we are seeing this issue most likely because we are not specifying the reference annotation when using a3m. Is there a way to specify the reference annotation in the a3m case? If not, would it be possible to add that option so that the conversion is not needed?
Reproduction
Input a3m created by removing all columns where the query had a gap and also removing deletions (lower-case letters) in non-query sequences so that everything has exactly the same length (
input.a3m):Converted input Stockholm (
input.sto):We then run the following pairs of commands:
hmmbuild --amino output_a3m.hmm input.a3mhmmbuild --amino output_sto.hmm input.stohmmbuild --amino --fast output_fast_a3m.hmm input.a3mhmmbuild --amino --fast output_fast_sto.hmm input.stohmmbuild --amino --hand output_hand_a3m.hmm input.a3m- fails with "Error: build failed: Alignment input has no reference annotation line"hmmbuild --amino --hand output_hand_sto.hmm input.sto- worksThe respective pairs for cases 1 and 2 don't match. E.g. running
diff --side-by-side --color --suppress-common-lines output_{a3m,sto}.hmmwill produce:Happy to provide extra details!