Category:Team Red B

After binding to the DNA junction as a dimer, the two subunits of the T7 endonuclease 1 catalyze cleavages independently but rather simultaneously before either is released from the junction (within the lifetime of the protein-junction complex) (Figure 6 and text around it, and Discussion). Similar characteristics have been found in T4 endonuclease VII.

Figure 1 shows significant degradation of both double and single stranded DNA upon infection with T7. The researchers, therefore, mutated a number of genes, and found out that only the mutants in genes 1 or 3 did not induce an increased endonuclease activity. It is, furthermore, stated that the product of gene 1 (RNA polymerase factor) is needed for the transcription of most of the T7 genome. Mutants in gene 3 were still synthesizing all essential T7 proteins, only lacking the increased endonuclease activity.

Figure 1 shows endonuclease activity after T7 infection of E. coli B. The endonuclease activity was not seen after infection with a mutant in gene 3 (Table 1).

The enzyme was purified and tested against various sources DNA. Breaks in the DNA strands were found (3-4 breaks per single strand before double strand breakage (see Figure 3).

"This packaging process is initiated by recognition and endonucleolytic cleavage of viral concatemeric DNA. Concatemeric DNA, which consists of head-to-tail unit-length molecules, is generally produced via recombination [21] or rolling-circle replication [34, 37]. Next, the cleaved DNA end is linked to the portal vertex of the empty prohead through specific interactions between the terminase and the portal protein [19, 20, 38, 41]. Thus, a packaging motor is assembled, which drives directional translocation of DNA into the prohead, powered by the energy of ATP hydrolysis"

P03 - large subnit

"The single p03 peak in the chromatogram suggests that the recombinant PaP3 large terminase subunit exists in solution as a monomer, which is also the case for many other phages [13, 16]."

"these results establish that the PaP3 large terminase subunit possesses a specific endonucleolytic activity on the PaP3 cos sequence, while the small subunit has a stimulatory effect on this activity."

p03 - large subunit

"The single p03 peak in the chromatogram suggests that the recombinant PaP3 large terminase subunit exists in solution as a monomer, which is also the case for many other phages [13, 16]."

"these results establish that the PaP3 large terminase subunit possesses a specific endonucleolytic activity on the PaP3 cos sequence, while the small subunit has a stimulatory effect on this activity. These two subunits may act cooperatively on the cos site of multimeric replicating PaP3 DNA and introduce staggered nicks to generate the 20-base ssDNA cohesive ends of the mature phage genome."

"This packaging process is initiated by recognition and endonucleolytic cleavage of viral concatemeric DNA. Concatemeric DNA, which consists of head-to-tail unit-length molecules, is generally produced via recombination [21] or rolling-circle replication [34, 37]. Next, the cleaved DNA end is linked to the portal vertex of the empty prohead through specific interactions between the terminase and the portal protein [19, 20, 38, 41]. Thus, a packaging motor is assembled, which drives directional translocation of DNA into the prohead, powered by the energy of ATP hydrolysis."

The NCBI BlastP shows protein sequence similarity between protein encoded by the LD11_gp054 gene in Bacillus phage Riley and the protein encoded by the LysB4 gene (also called BCB4_0006) in B. cereus phage B4.

BlastP values are 100% Query cover, E value = 8e-179, and Identity = 95%. It has been demonstrated that the LysB4 protein (produced by the LysB4 gene) exhibits endopeptidase activity in B. cereus phage B4. Therefore it can be inferred that this homologous protein exhibits the same function in Bacillus phage Riley.

The NCBI BlastP shows protein sequence similarity between the protein encoded by the Spock_55 gene in Bacillus phage Spock and the protein encoded by the LysB4 gene (also called BCB4_0006) in B. cereus phage B4.

BlastP values are 100% Query cover, E value = 0.0, and Identity = 99%. It has been demonstrated that the LysB4 protein (produced by the LysB4 gene) exhibits endopeptidase activity in B. cereus phage B4. Therefore it can be inferred that this homologous protein exhibits the same function in Bacillus phage Spock.

The NCBI BlastP shows protein sequence similarity between the protein encoded by the BigBertha_57 gene in Bacillus phage BigBertha and the protein encoded by the LysB4 gene (also called BCB4_0006) in B. cereus phage B4.

BlastP values are 81% Query cover, E value = 4e-101, and Identity = 77%. It has been demonstrated that the LysB4 protein (produced by the LysB4 gene) exhibits endopeptidase activity in B. cereus phage B4. Therefore it can be inferred that this homologous protein exhibits the same function in Bacillus phage BigBertha.

The NCBI BlastP shows protein sequence similarity between the protein encoded by gene 59 in Bacillus phage Troll and the protein encoded by the LysB4 gene (also called BCB4_0006) in B. cereus phage B4.

BlastP values are 100% Query cover, E value = 8e-164, and Identity = 94%. It has been demonstrated that the LysB4 protein (produced by the LysB4 gene) exhibits endopeptidase activity in B. cereus phage B4. Therefore it can be inferred that this homologous protein exhibits the same function in Bacillus phage Troll.

Figure 4, data not shown.

figure 6 shows gene product specific antibody binding to the tip of the phage tail