SEA-PHAGES | DNA binding domain protein or amidotransferase

Link to this post \| posted 11 Apr, 2019 16:35
dtobiason	In two cluster EA1 phages that we are annotating, gp55 (pham 39995) has a strong HHPred hit with a PDB crystal structure of Mur family proteins that are described as amidotransferases in the corresponding publication. Only one annotated EA1 genome calls this pham a DNA binding domain protein in Blastp with no other relevant hits. Not clear about the evidence for a DNA binding domain protein. Any help is much appreciated.

Link to this post | posted 11 Apr, 2019 16:35

In two cluster EA1 phages that we are annotating, gp55 (pham 39995) has a strong HHPred hit with a PDB crystal structure of Mur family proteins that are described as amidotransferases in the corresponding publication. Only one annotated EA1 genome calls this pham a DNA binding domain protein in Blastp with no other relevant hits. Not clear about the evidence for a DNA binding domain protein. Any help is much appreciated.

Link to this post \| posted 11 Apr, 2019 17:42
debbie	Hi Deb, As I look at this more closely (I HHPred'd BeeBee8 and Velene), the hits are alignments to their N-terminals (the last 20-25 amino acids). The predominant hit is to zinc-finger domains. I think that is what our protein is hitting - a zinc finger domain. Without more information, we should label these as Hypothetical Proteins. FYI - the rationale to DNA binding domains is that I think the zinc fingers were a term coined to mean a structure that inevitably binds to DNA (or RNA) using zinc ions. I think Wikipedia does a nice job of describing the specific and general use of the term 'zinc finger' and why we have not adopted it (since it now holds such a generalized meaning). https://en.wikipedia.org/wiki/Zinc_finger Edited 11 Apr, 2019 18:50

Link to this post | posted 11 Apr, 2019 17:42

debbie

Hi Deb,
As I look at this more closely (I HHPred'd BeeBee8 and Velene), the hits are alignments to their N-terminals (the last 20-25 amino acids). The predominant hit is to zinc-finger domains. I think that is what our protein is hitting - a zinc finger domain. Without more information, we should label these as Hypothetical Proteins.

FYI - the rationale to DNA binding domains is that I think the zinc fingers were a term coined to mean a structure that inevitably binds to DNA (or RNA) using zinc ions. I think Wikipedia does a nice job of describing the specific and general use of the term 'zinc finger' and why we have not adopted it (since it now holds such a generalized meaning).
https://en.wikipedia.org/wiki/Zinc_finger

Edited 11 Apr, 2019 18:50

Link to this post \| posted 16 Apr, 2019 14:04
dtobiason	Just to follow up. Pecaan listed the top two hits as strong hits with Mur family proteins (85% coverage and 98% probability) on HHpred for this gp, but when we just ran it thru HHpred the traditional way those hits did not exist. We asked HHpred to rerun Den3's gp55 and now the HHpred results are different and the original top hits are not there. We have been using Pecaan for annotating this semester and are concerned that we now have to go back to recheck all of the genes. We are finalizing our annotations now so please advise. Does Pecaan have a glitch? Can we still transfer Pecaan data into DNAmaster for our final file?

Link to this post | posted 16 Apr, 2019 14:04

dtobiason

Just to follow up. Pecaan listed the top two hits as strong hits with Mur family proteins (85% coverage and 98% probability) on HHpred for this gp, but when we just ran it thru HHpred the traditional way those hits did not exist. We asked HHpred to rerun Den3's gp55 and now the HHpred results are different and the original top hits are not there. We have been using Pecaan for annotating this semester and are concerned that we now have to go back to recheck all of the genes. We are finalizing our annotations now so please advise. Does Pecaan have a glitch? Can we still transfer Pecaan data into DNAmaster for our final file?

Link to this post \| posted 16 Apr, 2019 15:17
debbie	We can ask Claire about the glitch. Yes, you can still transfer data from PECAAN into DNA Master for your final file. debbie

Link to this post \| posted 17 Apr, 2019 14:46
ClaireRinehart	Deborah and Debbie, Both Den3 and Velene were put into PECAAN at the end of February 2019. For PECAAN, we have to do the HHpred searches locally since the high demand was putting to much load on the online site. At the end of Feb. we updated the database for pdb70 and changed their hhblits database for multi sequence alignment to Uniclust30 from the old uniprot20. This put the PECAAN HHpred more inline with what was being generated from the online site. This was probably after your entry of Den3 and Velene. For some of the other phages we had notices some differences between the online and PECAAN runs. That is what prompted the changes that we made, as mentioned above. Since the databases that we pull from are dynamic, we have put the dates at the top of each of the PECAAN database results to inform users of when the material was last updated. If ever you have a question about currency of the data, just press the re-run button adjacent to the database header to get the latest updates. They should be the most relevant. To update all of the data for a phage that was entered long before annotation, you can also go to the top Admin menu and select the Phages option. You can then find your phage by typing it's name into the search box. Press the Edit button at the end of the entry and then select Reblast… to update the Phagesdb and NCBI BLAST results or Rerun Evidence for all genes to update the evidence for all databases. Note that this will uncheck the evidence boxes that may have been previously marked. I hope this helps explain a little about where we have been and where we are today with PECAAN. -enjoy! Claire

Link to this post | posted 17 Apr, 2019 14:46

ClaireRinehart

Deborah and Debbie,
Both Den3 and Velene were put into PECAAN at the end of February 2019.
For PECAAN, we have to do the HHpred searches locally since the high demand was putting to much load on the online site.
At the end of Feb. we updated the database for pdb70 and changed their hhblits database for multi sequence alignment to Uniclust30 from the old uniprot20. This put the PECAAN HHpred more inline with what was being generated from the online site. This was probably after your entry of Den3 and Velene. For some of the other phages we had notices some differences between the online and PECAAN runs. That is what prompted the changes that we made, as mentioned above.
Since the databases that we pull from are dynamic, we have put the dates at the top of each of the PECAAN database results to inform users of when the material was last updated. If ever you have a question about currency of the data, just press the re-run button adjacent to the database header to get the latest updates. They should be the most relevant. To update all of the data for a phage that was entered long before annotation, you can also go to the top Admin menu and select the Phages option. You can then find your phage by typing it's name into the search box. Press the Edit button at the end of the entry and then select Reblast… to update the Phagesdb and NCBI BLAST results or Rerun Evidence for all genes to update the evidence for all databases. Note that this will uncheck the evidence boxes that may have been previously marked.
I hope this helps explain a little about where we have been and where we are today with PECAAN.
-enjoy!
Claire

Link to this post \| posted 17 Apr, 2019 14:51
dtobiason	Thanks Claire- that makes sense and thanks for the info about rerunning the evidence for all genes. PECAAN is super useful!

Link to this post \| posted 17 Apr, 2019 20:50
debbie	Thanks Claire!

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DNA binding domain protein or amidotransferase