Link to this post | posted 09 Aug, 2017 16:30
dbolliva	I am looking at cluster J phages and there are instances where two holins have been called. Allowed? The structure is LysA-holin-LysB-holin.

Link to this post | posted 09 Aug, 2017 16:26

dbolliva

Kristen, I reviewed several B1's and found really strong HHpred results for pham 5142 being PLA2. Granted the e values are not awesome, but the hit comes out of the CDD. All three of the B1s I reviewed had similar HHpred results. Not sure why virpocalpse is giving different results, but there is support for the other ones called I think. Note that the PLA2 hit is actually not from M smeg but a plant version. Best, Dave 21Kb

Link to this post | posted 04 Aug, 2017 14:19
dbolliva	On the function list that the SMRT team has been using RNAseE endoribonuclease is an approved function already. I think Welkin and Debbie are working on a significant revision of the annotation guide so this will be in the new version.

Link to this post | posted 03 Aug, 2017 23:39

dbolliva

In reviewing the CDD and HHpred for the Squint_196 Pham 25322 there is evidence that this protein is like the T4 immunity protein that sits in the plasma membrane and prevents superinfection. The Squint protein has three transmembrane helices. Is it possible to call this as an immunity protien or as imm. Admittedly the HHpred is only about half of the protein, but the combination makes it seem like this is a reasonable call. 272Kb

Link to this post | posted 03 Aug, 2017 14:48

dbolliva

Karen, The 3133 Pham is equally suspect. The HHPred resulta are for both RepA and DnaB helicases and the CDD results include RecA and AAA domains for ATP binding. Since I had so many DNA helicase hits of both types I currently have it annotated as a DNA helicase without the type specified. I am looking at Squint right now. I am doing this in PECAAN so you should be able to go look at the results if you like. Best, Dave

Link to this post | posted 02 Aug, 2017 19:26

dbolliva

Pham 6811 (with only three members, all J) appears to have two enzyme functions in the same polypeptide. The amino terminal region has strong HHpred results for methyltransferases (probability 98%) and also hits in CDD. The C terminus has hits on galactosyltransferases and glycosyltransferases (probability 99.7% for both) and also has hits in the CDD though the CDD hit is to glycosyltransferases. Currrently there is no ability to call combined methyltransferase/glycosyltransferase but it seems very likely that this is the case here. Can this be added? I have linked a copy of the HHpred result 58Kb

Link to this post | posted 01 Aug, 2017 22:12
dbolliva	Hi Karen, I am just getting started on looking at the J phages, but I noticed that our phage Squint does not appear to have a homolog of Klein_2 but has the homologs of Baka_5 and Baka_6 right next to each other. Just supporting the excellent sleuthing Jackie did. Dave

Link to this post | posted 31 Jul, 2017 15:40

dbolliva

A quick search revealed a paper with the following abstract: Transcription. 2012 Jan-Feb;3(1):2-7. doi: 10.4161/trns.3.1.18917. Structural and functional aspects of winged-helix domains at the core of transcription initiation complexes. Teichmann M1, Dumay-Odelot H, Fribourg S. The winged helix (WH) domain is found in core components of transcription systems in eukaryotes and prokaryotes. It represents a sub-class of the helix-turn-helix motif. The WH domain participates in establishing protein-DNA and protein-protein-interactions. Here, we discuss possible explanations for the enrichment of this motif in transcription systems. It seems we could call this a helix-turn helix or a winged helix would be more specific.

Link to this post | posted 20 Jul, 2017 21:33

dbolliva

I am trying to figure out how to call members of this Pham. There are many different functions that have been ascribed to it. I think this is due to the many conserved domains found in the protein as well as the large size. I am attaching a screen shot of the CDD results from when I BLASTed it on the NCBI website. There is a region towards the N-terminus that has a domain similar to primase/polymerases that are mostly eukaryotic/archeal. There is AAA ATPase domain that is also in a region with similarity to the RepA helicase and this also has similarity to DNA repair enzyme radB and also gamma and Tau subunits of DNA polymerase III. In the HHPRED results attached as a screenshot it is clear that there are two regions with different hits. The N-terminal region shows similarity to the primase/polymerase. The other region has the hits to thelicase/AAA/repair proteins. In the structure of the primase polymerase from the plasmid pRN1 found in an archaebacterium, these activities are dependent on the same amino acid residues, suggesting that both of these activities may use the same active site (EMBO Journal 22:2516-2525, Lipps et al). In the archaebacterial plasmid protein the enzyme was shown to have four activities, ATPase, primase, DNA polymerase, and helicase. In the most recent paper I found about this system (Microbiology Open, 2014,Berkner at al. 3:688-701) the authors suggest that this replication protein recognizes the origin of replication, opens stem loop structures in the origin with the helicase activity and then primes and makes a short DNA strand to create a replication fork that the host polymerase can then recognize and replicate. It is now clear why there is a number of different calls made for members of this Pham. Maybe it would make sense to call this a multifunctinal replication protein? I know we don't like nonspecific names, but the alternative might be DNA polymerase/primase/helicase/ ATPase? Edited 20 Jul, 2017 21:34 164Kb 386Kb

Link to this post | posted 20 Jul, 2017 17:32

dbolliva

A number of B1 phages contain a gene that has been called as AlpA or AlpA like (Pham 30124). AlpA is a protein present in the E. coli prophage cp4-57 and overexpression of this protein leads to excision of the prophage. Uniprot lists the E. coli AlpA as a DNA binding protein with a helix turn helix motif. The papers about E. coli AlpA do not address directly whether this protein binds DNA but expression of AlpA leads to higher transcription levels of another prophage protein that is an excisionase. When the B1 homolog is run through the HTH prediction software, there is no significant result. Other HHPRED results for B1 phages include excisionases including TorI and Xis. Both Xis and TorI have winged helix structures for binding DNA. Other proteins with the winged helix that are very similar are repressors (MuR). Ideas on whether we should continue to use AlpA or start calling these as winged helix proteins?