SEA-PHAGES | All posts created by fogartym1

Link to this post \| posted 16 May, 2024 19:50
fogartym1	Similar question here - PotPie: Gene 6 start 4858 , some evidence for minor capsid protein but unsure whether to call: The function list states ‘ "capsid morphogenesis protein" sometimes the CMP hits to ClpP proteases. If so, look for a serine-type endopeptidase activity. A significant hit to the CMP of D29 and L5 is sufficient evidence. We have a HHPred hit to D29 but not L5. https://toolkit.tuebingen.mpg.de/jobs/capsid 195/275 aligned P = 100 % Identities: 17%, Similarity: 0.223. Leaning towards NKF based on the low amino acid similarities/ Thanks Marie

Link to this post | posted 16 May, 2024 19:50

Similar question here -
PotPie:

Gene 6 start 4858 , some evidence for minor capsid protein but unsure whether to call:
The function list states ‘
"capsid morphogenesis protein" sometimes the CMP hits to ClpP proteases. If so, look for a serine-type endopeptidase activity. A significant hit to the CMP of D29 and L5 is sufficient evidence.

We have a HHPred hit to D29 but not L5.
https://toolkit.tuebingen.mpg.de/jobs/capsid

195/275 aligned P = 100 % Identities: 17%, Similarity: 0.223.
Leaning towards NKF based on the low amino acid similarities/
Thanks
Marie

Posted in: Annotation → capsid maturation protease or hypothetical protein (MuF-like minor capsid protein)

Link to this post \| posted 23 Jun, 2023 20:37
fogartym1	fogartym1 There are three DZ cluster phages in addition to Morgana. In each of the three DZ phages, there is a large gene of 2622 bp annotated as ADP-ribosyltransferase In Morgana, in the same area, there are two smaller genes encoded in two different frames - gp 141 ( ~ 80 bp) and gp 142. While BLAST gives hits for ADP-ribosyltransferase for both genes, only the second gene has a HHPRED hit and also 2 conserved domain hits from CDD for ADP-ribosyltransferses. I think that there is likely a frameshift in gp 141 (since there are no 1:1 matches for gp 42) and that is why there are two separate ORFs. I know that we cannot call a frameshift, but what should we do in terms of functional annotation? I am leaning towards calling gp141 as NKF and GP 142 as ADP-ribosyltransferase. I have attached the HHPRED hit for the ADP-ribosyltransferase hit. 58Kb

Link to this post | posted 23 Jun, 2023 20:37

fogartym1

fogartym1
There are three DZ cluster phages in addition to Morgana. In each of the three DZ phages, there is a large gene of 2622 bp annotated as ADP-ribosyltransferase

In Morgana, in the same area, there are two smaller genes encoded in two different frames - gp 141 ( ~ 80 bp) and gp 142. While BLAST gives hits for ADP-ribosyltransferase for both genes, only the second gene has a HHPRED hit and also 2 conserved domain hits from CDD for ADP-ribosyltransferses. I think that there is likely a frameshift in gp 141 (since there are no 1:1 matches for gp 42) and that is why there are two separate ORFs.
I know that we cannot call a frameshift, but what should we do in terms of functional annotation? I am leaning towards calling gp141 as NKF and GP 142 as ADP-ribosyltransferase.
I have attached the HHPRED hit for the ADP-ribosyltransferase hit.

Posted in: Annotation → Calling an ADP riboosyltransferase in DZ phage Morgana

Link to this post \| posted 23 Jun, 2023 20:32
fogartym1	There are three DZ cluster phages in addition to Morgana. In each of the three DZ phages, there is a large gene of 2622 bp annotated as ADP-ribosyltransferase In Morgana, in the same area, there are two smaller genes encoded in two different frames - gp 141 ( ~ 80 bp) and gp 142. While BLAST gives hits for ADP-ribosyltransferase for both genes, only the second gene has a HHPRED hit and also 2 conserved domain hits from CDD for ADP-ribosyltransferses. I think that there is likely a frameshift in gp 141 (since there are no 1:1 matches for gp 42) and that is why there are two separate ORFs. I know that we cannot call a frameshift, but what should we do in terms of functional annotation? I am leaning towards calling gp141 as NKF and GP 142 as ADP-ribosyltransferase. I have attached the HHPRED hit for the ADP-ribosyltransferase hit. 93Kb

Link to this post | posted 23 Jun, 2023 20:32

fogartym1

There are three DZ cluster phages in addition to Morgana. In each of the three DZ phages, there is a large gene of 2622 bp annotated as ADP-ribosyltransferase

In Morgana, in the same area, there are two smaller genes encoded in two different frames - gp 141 ( ~ 80 bp) and gp 142. While BLAST gives hits for ADP-ribosyltransferase for both genes, only the second gene has a HHPRED hit and also 2 conserved domain hits from CDD for ADP-ribosyltransferses. I think that there is likely a frameshift in gp 141 (since there are no 1:1 matches for gp 42) and that is why there are two separate ORFs.
I know that we cannot call a frameshift, but what should we do in terms of functional annotation? I am leaning towards calling gp141 as NKF and GP 142 as ADP-ribosyltransferase.
I have attached the HHPRED hit for the ADP-ribosyltransferase hit.

Posted in: Annotation → Calling an ADP riboosyltransferase in DZ phage Morgana

Link to this post \| posted 23 May, 2023 12:26
fogartym1	debbie Hi Marie, I think we have enough evidence to call this the capsid maturation protease (CMP). It hits a structural protein of a lactobacillus phage. Which then makes debate whether this is a head or tail protein. The next hit is to gp7 of SPP1 - the papers written about that gene are capsid proteins involved in packaging….. that makes me think not quite in the final virion structure…. And then we hit the capsid maturation protease of D29 (gp15). That hit is at 97%, with an e-value of 10-3. So while that hit is not the best, it is the most descriptive for me and jives with the stronger hits. AND then look at where this gene is in the genome. In between portal and scaffolding!! I think I would be OK calling is a CMP. what do you think? debbie Hi Debbie: Yes, this makes sense. The D29 hit to capsid maturation protease along with the synteny does seem like enough evidence. Thanks Marie

Link to this post | posted 23 May, 2023 12:26

fogartym1

debbie
Hi Marie,
I think we have enough evidence to call this the capsid maturation protease (CMP). It hits a structural protein of a lactobacillus phage. Which then makes debate whether this is a head or tail protein. The next hit is to gp7 of SPP1 - the papers written about that gene are capsid proteins involved in packaging….. that makes me think not quite in the final virion structure….
And then we hit the capsid maturation protease of D29 (gp15). That hit is at 97%, with an e-value of 10-3. So while that hit is not the best, it is the most descriptive for me and jives with the stronger hits. AND then look at where this gene is in the genome. In between portal and scaffolding!! I think I would be OK calling is a CMP. what do you think?
debbie

Hi Debbie:

Yes, this makes sense. The D29 hit to capsid maturation protease along with the synteny does seem like enough evidence.
Thanks
Marie

Posted in: Cluster-Specific Annotation Tips → Cluster DZ

Link to this post \| posted 22 May, 2023 19:46
fogartym1	Question about gene 5 in a DZ cluster phage, Morgana - call as minor capsid protein? ? Pham: 81331 This gene comes directly after portal and has 2 HHPRED hits > 90 % for minor head/capsid protein including P = 99.87to GP7_BPSPP Minor head protein GP7 OS=Bacillus phage SPP1 OX=10724 GN=7 PE=1 SV=2. It has been shown to be a minor capsid protein in SPP1. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6316719/ This gene was called as a Muf-like minor capsid protein in DZ phage Cafasso, and a minor capsid protein in phage Aleemily and some others. I know to discount the Muf, but is it appropriate to call as minor capsid protein in this area. I don't see any evidence for capsid maturation protease except this is where one would expect it based on synteny. HHPred image attached 221Kb

Link to this post | posted 22 May, 2023 19:46

fogartym1

Question about gene 5 in a DZ cluster phage, Morgana - call as minor capsid protein? ?
Pham: 81331

This gene comes directly after portal and has 2 HHPRED hits > 90 % for minor head/capsid protein including
P = 99.87to GP7_BPSPP Minor head protein GP7 OS=Bacillus phage SPP1 OX=10724 GN=7 PE=1 SV=2. It has been shown to be a minor capsid protein in SPP1. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6316719/
This gene was called as a Muf-like minor capsid protein in DZ phage Cafasso, and a minor capsid protein in phage Aleemily and some others. I know to discount the Muf, but is it appropriate to call as minor capsid protein in this area. I don't see any evidence for capsid maturation protease except this is where one would expect it based on synteny.

HHPred image attached

Posted in: Cluster-Specific Annotation Tips → Cluster DZ

Link to this post \| posted 13 May, 2020 15:44
fogartym1	I didn't manage to solve this issue, I addressed it in my cover letter and did my best with what was available, sorry!

Posted in: PECAAN → Rerun function not updating

Link to this post \| posted 25 Sep, 2019 12:39
fogartym1	Thanks Vic! We will try it out and report back on how it goes. Marie

Posted in: Phage Discovery/Isolation → Protocol to grow phage in culture?

Link to this post \| posted 22 Sep, 2019 18:10
fogartym1	Does anyone have a general protocol for growing phage in culture? We need to increase titers for a few of our phage and would love a protocol to get us started. Thanks

Posted in: Phage Discovery/Isolation → Protocol to grow phage in culture?

Link to this post \| posted 09 May, 2019 13:46
fogartym1	There are some recent published genome additions to Genbank and phagesdb that are very relevant for my annotation of Sixama. However, when I hit the rerun button for phagesdb and NCBI in PECAAN it is not updating. If I change the start site it updates, but for most genes I don't want to change the start site. Is there a way to rerun the databases (ideally for the entire genome at once)? Thanks Marie

Link to this post | posted 09 May, 2019 13:46

fogartym1

There are some recent published genome additions to Genbank and phagesdb that are very relevant for my annotation of Sixama. However, when I hit the rerun button for phagesdb and NCBI in PECAAN it is not updating. If I change the start site it updates, but for most genes I don't want to change the start site. Is there a way to rerun the databases (ideally for the entire genome at once)?
Thanks
Marie

Posted in: PECAAN → Rerun function not updating

Link to this post \| posted 24 May, 2018 02:05
fogartym1	Hi Welkin - This article is helpful, thank you. Based on this article, I would like to call Octobien14_gp36 as lysin A, N-acetyl-β-D-muramidase" - it has conserved domain pfam01510 as does phage BPs and BXZ1. It shows a 164 blast p score alignment (64 % query coverage) with lysin A of BPs. For Octobien_gp35, I would call it as lysin A, peptidase as does Supersulley mentioned in the first post above. Like Supersulley, it has conserved domain pfam 13529 and when blasted against Supersulley's lysin A, it has a score alignment of 129 (81% query coverage). Does this seem reasonable? Thanks Marie

Link to this post | posted 24 May, 2018 02:05

fogartym1

Hi Welkin - This article is helpful, thank you.

Based on this article, I would like to call Octobien14_gp36 as lysin A, N-acetyl-β-D-muramidase" - it has conserved domain pfam01510 as does phage BPs and BXZ1. It shows a 164 blast p score alignment (64 % query coverage) with lysin A of BPs.

For Octobien_gp35, I would call it as lysin A, peptidase as does Supersulley mentioned in the first post above.
Like Supersulley, it has conserved domain pfam 13529 and when blasted against Supersulley's lysin A, it has a score alignment of 129 (81% query coverage).

Does this seem reasonable?
Thanks
Marie

Posted in: Functional Annotation → Lysins in Gordonia

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