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I have to answer a question about a sub-task of my thesis: Can the bacteria X produce the protein Y?

I searched of course Google and BLAST. There is no data supporting that this specific bacteria ever produced this protein sequence.

However, my thesis supervisor wants me to go deeper[1], but I don't know how to do it in silico in a way to offer a proper answer.

What I'm thinking of doing with the data at hand is to create a DB on localhost with all the versions of the protein listed in BLAST (I'm using ruby, not sure how can I fetch all this data in FASTA format yet, but I'll figure it out), and all the strains of the bacteria listed in BLAST and perform a fuzzy string matching.

That's like brute-forcing. After I have a possible score I can get the closest score of them all, study it a bit and then submit my answer.

Is there any other way to attack this problem in order to submit a proper, at least partially polished answer?

NOTE: I don't have access to any lab, any experiment must be performed in silico.

[1] My supervisor believes that IF this bacteria needed this protein it WOULD be able to to produce it at will without undergoing mutations. And since his level of dedication and knowledge revolving molecular biology is scary, it's kind of hard to believe that he is wrong.

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I am coming from the wet lab - and my experience is that you can clone practically every gene into a fitting vector and express is in bacteria. You need the right regulatory sequences (promotors and so on), but it is possible. The question, if this protein is produced in big amount and if it soluble is something different. Also many proteins are modified posttranslationally which is something bacteria cannot do. –  Chris Jul 14 at 6:12
    
Hello Chris. I'm not interested in to cloning anything but thanks for the info. –  atmosx Jul 14 at 6:20
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I understood that. I simply wanted to add some comment on the practical side of this problem, this might help you as well? –  Chris Jul 14 at 6:31
    
Of course @Chris, I will mention to my paper that it's possible to achieve the wanted result with cloning. Without your comment I wouldn't had that extra paragraph, which will make my paper better anyway :-) ... So thank you again! –  atmosx Jul 14 at 7:04
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you can predict an ORF from DNA/RNA sequence but you cannot be sure if the protein is produced. You can either do LC-MS to see if signature peptides are produced or use molecular bio experiments as Chris suggested. You can also tag your sequence with GFP and see if it expresses. –  WYSIWYG Jul 17 at 9:01

1 Answer 1

up vote 1 down vote accepted

Your problem will finally boil down to searching your sequence in the Blast databases. Performing Blast seems to be probably the best way to find out if your bacteria has that specific protein expressed or not. If you could not find it in the nearest species using Blast, then try running PSI-BLAST, which would return you distant homologs, by which you can see if your protein is expressed in the other bacterial species.

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I'm on to something using blast (I dropped out the idea of writing my own script to do this as soon as I played around with BLAST). I know that E. Coli can produce my protein (it's documented) at protein level. I couldn't find a perfect match thought, I've found a 74% positive match. Then searched about Desulfovibrio and found a 75% positive match (for a specific strain). I think it's enough, I can't take the risk to answer the question in a definitive way but, it's enough data to give some food for thought and possible direction. Comments are more than welcomed! Ty! –  atmosx Jul 14 at 11:42

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