PMID:26873847

Citation	Song, J, Xia, F, Jiang, H, Li, X, Hu, L, Gong, P, Lei, L, Feng, X, Sun, C, Gu, J and Han, W (2016) Identification and characterization of HolGH15: the holin of Staphylococcus aureus bacteriophage GH15. J. Gen. Virol. 97:1272-81
Abstract	Holins are phage-encoded hydrophobic membrane proteins that spontaneously and non-specifically accumulate and form lesions in the cytoplasmic membrane. The ORF72 gene (also designated HolGH15) derived from the genome of the Staphylococcus aureus phage GH15 was predicted to encode a membrane protein. An analysis indicated that the protein encoded by HolGH15 potentially consisted of two hydrophobic transmembrane helices. This protein exhibited the structural characteristics of class II holins and belonged to the phage_holin_1 superfamily. Expression of HolGH15 in Escherichia coli BL21 cells resulted in growth retardation of the host cells, which was triggered prematurely by the addition of 2,4-dinitrophenol. The expression of HolGH15 caused morphological alterations in engineered E. coli cells, including loss of the cell wall and cytoplasmic membrane integrity and release of intracellular components, which were visualized by transmission electron microscopy. HolGH15 exerted efficient antibacterial activity at 37 °C and pH 5.2. Mutation analysis indicated that the two transmembrane domains of HolGH15 were indispensable for the activity of the full-length protein. HolGH15 showed a broad antibacterial range: it not only inhibited Staphylococcus aureus, but also demonstrated antibacterial activity against other species, including Listeria monocytogenes, Bacillus subtilis, Pseudomonas aeruginosa, Klebsiella pneumoniae and E. coli. At the minimal inhibitory concentration, HolGH15 evoked the release of cellular contents and resulted in the shrinkage and death of Staphylococcus aureus and Listeria monocytogenes cells. To the best of our knowledge, this study is the first report of a Staphylococcus aureus phage holin that exerts antibacterial activity against heterogeneous pathogens.
Links	PubMed Online version:10.1099/jgv.0.000428
Keywords
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
9CAUD:I6P9Q0		GO:0019835: cytolysis	ECO:0000314:		P		Figure 6 shows the cytolysis breakdown of the cell membrane and cell walls when bacterial host cells are exposed to a vector carrying the HolGH15 gene.	complete
9CAUD:I6P9Q0		GO:0016021: integral component of membrane	ECO:0000314:		C		Figure 3d shows the expression of HolGH15 over 10 minute intervals after western blotting was performed. The E. coli BL21-CodonPlus cells harbouring pET-28a-HOLGH15 were induced with IPTG to detect HolGH15. The result demonstrated the presence of a band of about 21 kDa with consistent size with the predicted mass of HolGH15. The band was detected 10 minutes after the induction with IPTG. Figure 3b: Transmission Electron Microscopy observations showed damage and disruption in the induced E. coli cells. The result was the release of cellular contents and showed apperance simillar to empty, non-living bacterial envelopes of Gram-negative bacteria.	complete CACAO 12665

References

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