SEA-PHAGES | All posts created by NWCiowaSEAPHAGE

Link to this post \| posted 06 Oct, 2024 00:55
NWCiowaSEAPHAGE	We're looking for protocols and reagents folks have used successfully for TEM that are not radioactive. Thanks in advance for any help you can give us!

Posted in: Phage Discovery/Isolation → Non-radioactive TEM stain

Link to this post \| posted 03 Jun, 2023 15:30
NWCiowaSEAPHAGE	We recently annotated two EM phages. The auto-annotations for both produced many many false positive genes. We saw reverse genes without BLAST hits without much coding potential completely overlapping forward genes with good CP and BLAST hits. We had to delete nearly half of the auto-annotated genes. Debbie added: Note when you review this in DNA maser, you will find the GeneMark has called all of the forward genes and Glimmer has called all of the reverse genes. this is based on how these 2 programs call this differently OR is it because there is a big open reading frame in the same location and the programs discriminate which one to pick differently. If you simply discard one of the sets of data, are you discarding possible genes. if all of these genes are in PECAAN, always good to see if you are throwing away anything with a functional…. Edited 06 Jun, 2023 02:05 521Kb

Link to this post | posted 03 Jun, 2023 15:30

NWCiowaSEAPHAGE

We recently annotated two EM phages. The auto-annotations for both produced many many false positive genes. We saw reverse genes without BLAST hits without much coding potential completely overlapping forward genes with good CP and BLAST hits. We had to delete nearly half of the auto-annotated genes.

Debbie added: Note when you review this in DNA maser, you will find the GeneMark has called all of the forward genes and Glimmer has called all of the reverse genes. this is based on how these 2 programs call this differently OR is it because there is a big open reading frame in the same location and the programs discriminate which one to pick differently. If you simply discard one of the sets of data, are you discarding possible genes. if all of these genes are in PECAAN, always good to see if you are throwing away anything with a functional….

Edited 06 Jun, 2023 02:05

Posted in: Cluster EM Annotation Tips → Putative Genes on Both Strans in the same location

Link to this post \| posted 14 Mar, 2022 15:10
NWCiowaSEAPHAGE	Thanks! Very helpful!

Posted in: Functional Annotation → SecB-like translocase or SecB export protein??

Link to this post \| posted 04 Mar, 2022 16:25
NWCiowaSEAPHAGE	Mitch, Sorry! The numbering has changed since we annotated. Here is its sequence: MPVTPGDVTGARIRWRERSAVRHYDGDPEPSYREDVDEEDGPFKLGAEIGRGTDDDDTPIFSVRMKGSFTKPDAQVRIDVEVVFKIDSGEEVDQAFIQKHAMPYVFGYVRGGFTDACRSVGLPGWMIPMVDLAADIQYEDA - sara

Posted in: Functional Annotation → SecB-like translocase or SecB export protein??

Link to this post \| posted 04 Mar, 2022 14:57
NWCiowaSEAPHAGE	balish This protein certainly looks like a relative of SecB, whose actual function is as a chaperone in protein secretion that delivers its unfolded substrates to SecA. Looking quickly at the paper on the structure of SecB from Haemophilus influenzae, I note that the phage protein doesn't conserve all the residues known to be important for the SecB-SecA interaction, so I don't think it's likely that it does what SecB does. Whether it could be a chaperone at all is a good question, and a more difficult one. If anything, I might consider calling it a SecB-related protein, but I'm not too comfortable going beyond that. Hi Mitch! I'm getting HHPred hits from Wrigley_46 to SecB proteins too. The hits are good (probability = 99.8 with 96% coverage). I looked carefully at the sequence of Wrigley_46 and see a D at position 25 (not 20), an E at position 29 (not 24), an I (not L but conservative substitution) at position 78 (not 75) and an E at position 81 (not 77). The Randall et al paper from 2004 also indicates the importance of a C-terminal helix which I have in Wrigley_46. Any thoughts about this one being SecB-like? Enough to justify a functional call or not? I do see in Sala et al 2014 that Mycobacterium have a SecB-like protein (Wrigley is a Gordonia phage). Thanks for your (and anyone else's) help! - sara Edited 04 Mar, 2022 14:58

Link to this post | posted 04 Mar, 2022 14:57

NWCiowaSEAPHAGE

balish
This protein certainly looks like a relative of SecB, whose actual function is as a chaperone in protein secretion that delivers its unfolded substrates to SecA. Looking quickly at the paper on the structure of SecB from Haemophilus influenzae, I note that the phage protein doesn't conserve all the residues known to be important for the SecB-SecA interaction, so I don't think it's likely that it does what SecB does. Whether it could be a chaperone at all is a good question, and a more difficult one. If anything, I might consider calling it a SecB-related protein, but I'm not too comfortable going beyond that.

Hi Mitch! I'm getting HHPred hits from Wrigley_46 to SecB proteins too. The hits are good (probability = 99.8 with 96% coverage). I looked carefully at the sequence of Wrigley_46 and see a D at position 25 (not 20), an E at position 29 (not 24), an I (not L but conservative substitution) at position 78 (not 75) and an E at position 81 (not 77). The Randall et al paper from 2004 also indicates the importance of a C-terminal helix which I have in Wrigley_46. Any thoughts about this one being SecB-like? Enough to justify a functional call or not? I do see in Sala et al 2014 that Mycobacterium have a SecB-like protein (Wrigley is a Gordonia phage).
Thanks for your (and anyone else's) help!
- sara

Edited 04 Mar, 2022 14:58

Posted in: Functional Annotation → SecB-like translocase or SecB export protein??

Link to this post \| posted 15 Feb, 2022 16:06
NWCiowaSEAPHAGE	lhughes Another one from Abt2graduatex2: RNase adapter protein RapZ The genome has this function (not on the approved list) for gp55. I compared the HHPRED data for the equivalent gene in BabyGotBac (gp56, which was annotated as "ATPase" and found there is 100% probability, 90% coverage, and E-value of 0 to the RNase adapter protein (RapZ in E.coli). The function in Uniprot is given as "Modulates the synthesis of GlmS, by affecting the processing and stability of the regulatory small RNA GlmZ" Thoughts on this function? Lee Hi! I'm looking to see if Lee got an answer on this one and can't find it. We have phage Santhid (cluster DY) gp56 with a similar hit on HHPred. 100% probability 86% coverage e = 1x10-32. The hit is to a RNAse adapter protein RapZ. What was your final call and why, Lee? Thanks, Sara

Link to this post | posted 15 Feb, 2022 16:06

NWCiowaSEAPHAGE

lhughes
Another one from Abt2graduatex2: RNase adapter protein RapZ

The genome has this function (not on the approved list) for gp55. I compared the HHPRED data for the equivalent gene in BabyGotBac (gp56, which was annotated as "ATPase" and found there is 100% probability, 90% coverage, and E-value of 0 to the RNase adapter protein (RapZ in E.coli).

The function in Uniprot is given as "Modulates the synthesis of GlmS, by affecting the processing and stability of the regulatory small RNA GlmZ"

Thoughts on this function?

Lee

Hi! I'm looking to see if Lee got an answer on this one and can't find it. We have phage Santhid (cluster DY) gp56 with a similar hit on HHPred. 100% probability 86% coverage e = 1x10-32. The hit is to a RNAse adapter protein RapZ. What was your final call and why, Lee?
Thanks, Sara

Posted in: Functional Annotation → Functions not on the approved list

Link to this post \| posted 16 Jun, 2021 16:26
NWCiowaSEAPHAGE	I had a hard time finding this link when I needed it last so I'm putting it here for me later and for any of you who might also need an easy place to find it https://phagesdb.org/blog/posts/26/

Posted in: Sequencing, Assembling, and Finishing Genomes → Tagmentation

Link to this post \| posted 16 Mar, 2021 15:24
NWCiowaSEAPHAGE	We originally called it a capsid maturation protease but don't see a scaffolding protein so I thought, based on Welkin's message we should rename it. I'm good with capsid maturation protease. Maybe take this thread down. It confused us .

Posted in: Cluster B Annotation Tips → Capsid maturation protease and MuF-like fusion protein

Link to this post \| posted 16 Mar, 2021 14:07
NWCiowaSEAPHAGE	I don't think capsid morphogenesis protein is on the approved functions list. Could someone add it please? Thanks!

Posted in: Cluster B Annotation Tips → Capsid maturation protease and MuF-like fusion protein

Link to this post \| posted 12 Mar, 2021 18:49
NWCiowaSEAPHAGE	Thanks Chris! I will definitely work through that paper with my students.

Posted in: Annotation → Membrane proteins

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