Annotation using Pfam

Question

[Md Moinuddin]

Hi Community,

I have 17 metagenomically assembled genomes (MAGs). I want to annotate the protein of these genome using Pfam.

I want a step by step guideline for the annotations in Linux bash.

My idea is: I need to download the Pfam database but there are lot of files and I am confused which one should I download. Next I need to use HMM for the annotations.

Thank you so much!

Answer 1

[Mensur Dlakic]

This is not going to be step by step, but hopefully still useful. First download and unpack Pfam HMM database:

wget https://ftp.ebi.ac.uk/pub/databases/Pfam/current_release/Pfam-A.hmm.gz
gunzip Pfam-A.hmm.gz
hmmpress Pfam-A.hmm

Assuming your protein sequences are in proteome.faa, do the search:

hmmscan -o annotations.txt --tblout annotations.tbl -E 0.01 --cpu 4 Pfam-A.hmm proteome.faa

Programs from the HMMer package are described here.

[Md Moinuddin]

Thank you so much

[aldreaspect2008]

Dear Prof/Dr/Sir,

May I kindly ask, is the E-value used usually is 0.01? Or we can be more stringent with going with 1e-5 which are taught by the academician as a usual standards (to expect 1 error in 100,000)?

Thank you very much

[Mensur Dlakic]

I think you already answered your own question. If you want to be more stringent and get strong hits without any manual checking, go with lower E-values. If you want to include more potential hits for later manual inspection, higher E-values are acceptable.