Link to this post | posted 11 Aug, 2017 01:54

delesall

In reviewing the annotation of phage LilHazelnut, I noticed two genes, ORF 68, start 45763, and ORF 69, start 46743, with decent HHPred matches (98% prob, 40% coverage, reasonably low E-values) to respectively G38P, the N-terminal phage replisome organiser (The protein contains a region of low-complexity sequence that reflects DNA direct repeats able to function as an origin of phage replication) and G39P, Loader and inhibitor of phage G40P. G39P inhibits the initiation of DNA replication by blocking G40P replicative helicase. Although the protein of ORF 70 does not have matches to a replicative helicase, should we call some function for ORF 68 and 69, and if so, what? From Ayora et al (1999) J Mol Biol. 288(1):71-85) Abstract Initiation of Bacillus subtilis bacteriophage SPP1 replication requires the phage-encoded genes 38, 39 and 40 products (G38P, G39P and G40P). G39P, which does not bind DNA, interacts with the replisome organiser, G38P, in the absence of ATP and with the ATP-activated hexameric replication fork helicase, G40P. G38P, which specifically interacts with the phage replication origin (oriL) DNA, does not seem to form a stable complex with G40P in solution. G39P when complexed with G40P-ATP inactivates the single-stranded DNA binding, ATPase and unwinding activities of G40P, and such effects are reversed by increasing amounts of G38P. Unwinding of a forked substrate by G40P-ATP is increased about tenfold by the addition of G38P and G39P to the reaction mixture. The specific protein-protein interactions between oriL-bound G38P and the G39P-G40P-ATPgammaS complex are necessary for helicase delivery to the SPP1 replication origin. Formation of G38P-G39P heterodimers releases G40P-ATPgammaS from the unstable oriL-G38P-G39P-G40P-ATPgammaS intermediate. G40P-ATPgammaS binds to the origin region, the uncomplexed G38P fraction remains bound to oriL, and the G38P-G39P heterodimer is lost from the complex. We demonstrate that G39P is a component of an oligomeric nucleoprotein complex which plays an important role in the initiation of SPP1 replication.

Link to this post | posted 10 Aug, 2017 19:29

delesall

In reviewing the annotation of Demsculpinboyz (an F2),the protein of ORF 82 (start 43366, stop 44214) has blast matches to tRNA methyl transferase, PAPs reductase, and ribonucleotide reductase! There is no CDD hits but a number of good HHPred matches (Prob 99%, coverage greater than 65%, low E-values)to 1) phosphoadenosine phosphosulfate sulfurtransferase, 2) Adenine nucleotide alpha hydrolases superfamily, and 3) adenosine 5'-phosphosulfate reductase. All this suggests that going with ribonucleotide reductase is safest bet. What does the forum think?

Link to this post | posted 10 Aug, 2017 18:19
delesall	For the output, are we just looking for the absence or presence of a coiled coil motif?

Link to this post | posted 03 Jul, 2017 17:30
delesall	I would still want to call both genes - the "original" gene in other phages as no known function. The question is whether I annotate both as "hypothetical protein" or "hypothetical protein, N-terminus" and "hypothetical protein, C-terminus". And what Genbank will accept…

Link to this post | posted 16 Jun, 2017 20:30
delesall	Thanks Chris That does look like a possible answer to the problem. I think we may need to go back to BYU for the sequencing data!

Link to this post | posted 06 Jun, 2017 15:50
delesall	Yes - see attached file - 34B is (obviously) between labeled 34 and 35 9Kb

Link to this post | posted 06 Jun, 2017 15:38
delesall	From HHPred (as well as Interpro, pfam, … Ead/Ea22-like protein This family contains phage proteins and bacterial proteins that are likely to represent integrated phage proteins. This family includes the Lambda phage Ea22 early protein as well as the Bacteriophage P22 Ead protein. A quick Google search does not find more information than this… I'm happy to just say NKF

Link to this post | posted 02 Jun, 2017 21:26
delesall	Gene 51 in Smairt (RVS 39018-37300) has interesting function calls in phages db: mostly phosphoesterase but also RDF/NrdC. Conserved domain database shows Hedgehog/Intein domain, found in Hedgehog proteins as well as proteins which contain inteins and undergo protein splicing. So, I'm planning to go with Phosphoesterase with intein domain. Do I get a second?

Link to this post | posted 02 Jun, 2017 21:19

delesall

In Smairt I have two genes that overlap (ca 550 bp) that both have blastp matches to the same gene porduct in JC27. I have aligned the DNA sequences to that gene and get the following (see attached file as well): Smairt 34B aligns to JC27 gene 37 from base 1 to base 588 with a 11 snps and one indel. Smairt 35 aligns from base 143 to base 688 with 8 snps and one indel. I cannot make Smairt 35 longer to include the first 142 bp with matches in JC27 and 34B cannot be made longer (darn stop codon) to include the last 100 bp that match JC27… None of these putative ORFs have known functions. Call both ORF 34B and 35 in Smairt despite 550bp overlap or just call ORF 34B (which is longer than 35? That is my dimemma 2Kb

Link to this post | posted 02 Jun, 2017 20:58
delesall	I have a cluster A1 phage (Smairt) with an ORF with nice blastp and HHPred matches (98% probability, 70% coverage) to Ead/Ea22-like protein. Call function?