title: Identification and characterization of HolGH15: the holin of Staphylococcus aureus bacteriophage GH15.
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.
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Jun
authors.initials: J
authors.lastname: Song
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Feifei
authors.initials: F
authors.lastname: Xia
authors.affiliation: The First Affiliated Hospital to Changchun University of Chinese Medicine,Changchun 130021, PRChina.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Jiang
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Xinwei
authors.initials: X
authors.lastname: Li
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Liyuan
authors.initials: L
authors.lastname: Hu
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Pengjuan
authors.initials: P
authors.lastname: Gong
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Liancheng
authors.initials: L
authors.lastname: Lei
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Xin
authors.initials: X
authors.lastname: Feng
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Changjiang
authors.initials: C
authors.lastname: Sun
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.
authors.forename: Jingmin
authors.initials: J
authors.lastname: Gu
authors.affiliation: College of Veterinary Medicine,Jilin University, Changchun 130062, PRChina.|Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonoses,Yangzhou 225009, PRChina.
authors.forename: Wenyu
authors.initials: W
authors.lastname: Han
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents; D003001:Cloning, Molecular; D003593:Cytoplasm; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D008089:Listeria monocytogenes; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D013204:Staphylococcus Phages / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000821:virology*; D000066608:Underage Drinking; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D008565:Membrane Proteins; D014764:Viral Proteins