Gene 2
Phage Name: 
Stop Coordinate 
Direction (Forward or Reverse) 
Gap (Overlap) with Previous Gene 
106 bp gap 
Selected Start Coordinate 
Selected Function 
Hypothetical Protein 
Annotation Decision 1 (Is this a gene?): 
Gathering Evidence 
Explain Your Rationale 
Was the gene called by an auto-annotation program (Glimmer, GeneMark) 
Glimmer call at 733 
Genemark call at 718 
This provides evidence that the start site can be at either position, but evidences that this is a gene. 
"Original Glimmer call @bp 733 has strength 8.69; GeneMark calls start at 718" 
Is there evidence for coding potential? 
Yes, coding potential in GeneMarkS is visible. This is pictured below. GeneMark also appears to display coding potential as well.  
Is the gene present in other annotated genomes? 
Yes, phamerator matches the Lewando gene 2 with homologous genes in phages Zeina and Uzumaki for gene 2.  
Does the gene violate any of the major guiding principles? 
Yes, this is a gene 
Annotation Decision 2 (What is the best possible start site for this gene?): 
Gathering Evidence 
Explain Your Rationale 
What start site do Glimmer and GeneMark suggest? 
Glimmer call at 733, GeneMark call at 718 
"Original Glimmer call @bp 733 has strength 8.69; GeneMark calls start at 718" 
Does the start site have an associated ribosome binding site with a high score? 
Yes. There is Z score of 2.865 for the start at 733, best of all possible sites. The 733 site has the highest total score as well and the distance is acceptable.  
Is the predicted start codon the longest ORF? If not, does the longest ORF result in excessive overlap (>30bp)? 
No, and there will be no excessive overlap if a upstream start site is selected. This shows that this region does not maximize coding potential - and suggests that the start site may be further upstream.  
Is the start site conserved in other genomes as indicated by Starterator? 
Yes, conservation in the start site. 37 other phages share this same start site - indicating that the 733bp start site may be a conserved start site 
Is the start site conserved in other phage genomes as indicated by BLASTp? 
Yes, 100% query cover with another Zeina protein (Zeina_2) given 733 start site. Through a homologous start site, the 733 start site is further supported. 
BlastP PhagesDB: Q1:S1 
BlastP NCBI: Q1:S1 
BlastP DNA Master: Q1:1 
733 is the start site 
Annotation Decision 3 (What is the function of the putative protein?): 
Gathering Evidence 
Explain Your Rationale 
Does the protein align with a protein having functional assignment in BLASTp (phagesDB and/or GenBank) with an alignment of 10^-4 or smaller? 
No, there are high scoring matches, but unfortunately, there is no functional assignment given to these proteins. The most similair protein is the hypotheitical protein Zein_2. E value for phages db - 1e-56, E value for NCBI - 4e-73, e-value for DNA master - 0  
PHAGES DB: 
Does this protein align with a protein having a functional assignment in the PDB or other database in HHPred with a probability of 90% or greater with appropriate coverage? 
No quality match. 
No, there is no alignment of high order, with probabilities falling far below the 90% mark. It appears that this gene does contain a urease domain though, as indicated in the image below. Top match in PDB/NCBI database.  
Is the gene located adjacent to genes of known function and in a region of the genome that shows high conservation of gene order? 
Yes, this gene is located in a synteic region of the phage genome. The next two genes are conserved throughout the AU6 subclusterr and the same preceding gene is peresent in Uzumaki relative to Lewando. Proximal genes include HNH endonuclease and endolysin, two proteins which may be important in setting up the bacterial cell for infection.  
Is the proposed function found on the SEA-PHAGES approved function list? 
Hypothetical Protein 