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Brok-Volchanskaya, VS, Kadyrov, FA, Sivogrivov, DE, Kolosov, PM, Sokolov, AS, Shlyapnikov, MG, Kryukov, VM and Granovsky, IE (2008) Phage T4 SegB protein is a homing endonuclease required for the preferred inheritance of T4 tRNA gene region occurring in co-infection with a related phage. Nucleic Acids Res. 36:2094-105


Homing endonucleases initiate nonreciprocal transfer of DNA segments containing their own genes and the flanking sequences by cleaving the recipient DNA. Bacteriophage T4 segB gene, which is located in a cluster of tRNA genes, encodes a protein of unknown function, homologous to homing endonucleases of the GIY-YIG family. We demonstrate that SegB protein is a site-specific endonuclease, which produces mostly 3' 2-nt protruding ends at its DNA cleavage site. Analysis of SegB cleavage sites suggests that SegB recognizes a 27-bp sequence. It contains 11-bp conserved sequence, which corresponds to a conserved motif of tRNA TpsiC stem-loop, whereas the remainder of the recognition site is rather degenerate. T4-related phages T2L, RB1 and RB3 contain tRNA gene regions that are homologous to that of phage T4 but lack segB gene and several tRNA genes. In co-infections of phages T4 and T2L, segB gene is inherited with nearly 100% of efficiency. The preferred inheritance depends absolutely on the segB gene integrity and is accompanied by the loss of the T2L tRNA gene region markers. We suggest that SegB is a homing endonuclease that functions to ensure spreading of its own gene and the surrounding tRNA genes among T4-related phages.


PubMed PMC2330249 Online version:10.1093/nar/gkn053


Bacteriophage T4/enzymology; Bacteriophage T4/genetics; Base Sequence; Conserved Sequence; DNA, Viral/chemistry; DNA, Viral/metabolism; Endodeoxyribonucleases/genetics; Endodeoxyribonucleases/metabolism; Gene Conversion; Inheritance Patterns; Myoviridae/genetics; Open Reading Frames; RNA, Transfer/genetics; Substrate Specificity; T-Phages/genetics; Viral Proteins/genetics; Viral Proteins/metabolism



Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status


GO:0004520: endodeoxyribonuclease activity



The T4 SegB protein is a site-specific endonuclease that recognizes a 27-bp sequence. 666 bp segB ORF was placed under the control of T7 promoter in pET-15b vector. SegB protein was overproduced in E. coli T7 lac promoter system and purified. The purified protein was incubated with a linear plasmid DNA and products were analyzed of the reaction through gel electrophoresis (Figure 3). SegB cleaved DNA by introducing double-strand breaks in reactions requiring Mg2+, Mn2+, or Ca2+ cations. With Mg2+ and Ca2+ cation, SegB cleaved the DNA at 1 point, creating 2.2 kb and 1.6 kb fragments. In the presence of Mn2+, multiple cleavage sites were present.

CACAO 13238


GO:0004519: endonuclease activity



Bacteriophage T4 segB gene encodes for a protein of unknown function, but is homologous to homing endonucleases of the GIY-YIG family. By using sequence alignment with PCR, the study demonstrates that the SegB protein is a site-specific endonuclease, by analyszing the DNA cleavage site. The cleavage site analysis suggests SegB is a homing endonuclease that functions to spread its own gene among T4-related phages. In Figure 3, it is shown that Phage T4 SegB protein demonstrates endonuclease activity by cleaving DNA in the presence of Mg2+, Mn2+ or Ca2+ cations. Although the study uses a wet lab, the focus is on sequencing; the evidence is sequence alignment, instead of direct assay.

Missing: with/from
CACAO 13299


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