Restriction and modification of deoxyarchaeosine (dG + )-containing phage 9 g DNA

• Published in: Scientific reports Vol. 7; no. 1; pp. 8348 - 13
• Main Authors: Tsai, Rebecca, Corrêa, Ivan R, Xu, Michael Y, Xu, Shuang-Yong
• Format: Journal Article
• Language: English
• Published: England Nature Publishing Group 21.08.2017; Nature Publishing Group UK; Nature Portfolio
• Subjects: Adenine; Coliphages - genetics; Cytosine; Deoxyribonucleases, Type II Site-Specific - metabolism; Deoxyribonucleic acid; DNA; DNA primase; DNA, Viral - chemistry; DNA-directed DNA polymerase; E coli; Escherichia coli - genetics; Escherichia coli - growth & development; Gene Expression Regulation, Viral; Genomes; Glutamine; Glutamine amidotransferase; Guanosine - analogs & derivatives; Guanosine - chemistry; Guanosine - genetics; Liquid chromatography; Mass spectroscopy; Methylation; Methyltransferases - metabolism; Phages; Primase; Terminase
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-017-08864-4

E. coli phage 9 g contains the modified base deoxyarchaeosine (dG ) in its genome. The phage encodes its own primase, DNA ligase, DNA polymerase, and enzymes necessary to synthesize and incorporate dG . Here we report phage 9 g DNA sensitivity to >200 Type II restriction endonucleases (REases). Among the REases tested approximately 29% generated complete or partial digestions, while the remaining 71% displayed resistance to restriction. Phage 9 g restriction fragments can be degraded by DNA exonucleases or ligated by T3 and T4 DNA ligases. In addition, we examined a number of cytosine and adenine methyltransferases to generate double base modifications. M.AluI, M.CviPI, M.HhaI, and M.EcoGII were able to introduce C or A into 9 g DNA as confirmed by partial resistance to restriction and by liquid chromatography-mass spectrometry. A number of wild-type E. coli bacteria restricted phage 9 g, indicating natural restriction barriers exist in some strains. A BlastP search of GenBank sequences revealed five glutamine amidotransferase-QueC homologs in Enterobacteria and Pseudomonas phage, and distant homologs in other phage and bacterial genomes, suggesting that dG is not a rare modification. We also mapped phage 9 g DNA packaging (pac) site containing two 21-bp direct repeats and a major terminase cleavage site in the phage genome.