World J Virol. 2013 Nov 12;2(4):152-9. doi: 10.5501/wjv.v2.i4.152.

Genetic analysis of structural proteins in the adsorption apparatus of bacteriophage epsilon 15.

World journal of virology

Jared A Guichard, Paula C Middleton, Michael R McConnell

PMID: 24286036 PMCID: PMC3832910 DOI: 10.5501/wjv.v2.i4.152

Abstract

AIM: To probe the organizational structure of the adsorption apparatus of bacteriophage epsilon 15 (E15) using genetic and biochemical methodology

METHODS: Hydroxylamine was used to create nonsense mutants of bacteriophage E15. The mutants were then screened for defects in their adsorption apparatus proteins, initially by measuring the concentrations of free tail spike proteins in lysates of cells that had been infected by the phage mutants under non-permissive growth conditions. Phage strains whose infected cell lysates contained above-average levels of free tail spike protein under non-permissive growth conditions were assumed to contain nonsense mutations in genes coding for adsorption apparatus proteins. These mutants were characterized by classical genetic mapping methods as well as automated sequencing of several of their genes. Finally, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography were used to examine the protein compositions of the radioactive particles produced when the various mutants were grown on a non-permissive host cell in the presence of (35)S-methionine and co-purified along with E15wt phage on CsCl block gradients.

RESULTS: Our results are consistent with gp4 forming the portal ring structure of E15. In addition, they show that proteins gp15 and gp17 likely comprise the central tube portion of the E15 adsorption apparatus, with gp17 being more distally positioned than gp15 and dependent upon both gp15 and gp16 for its attachment. Finally, our data indicates that tail spike proteins comprised of gp20 can assemble onto nascent virions that contain gp7, gp10, gp4 and packaged DNA, but which lack both gp15 and gp17, thereby forming particles that are of sufficient stability to survive CsCl buoyant density centrifugation.

CONCLUSION: The portal ring (gp4) of E15 is bound to tail spikes (gp20) and the tail tube (gp15 and gp17); gp17's attachment requires both gp15 and gp16.

Keywords: Epsilon15; Salmonella phages; Virion structure

References

1. Virology. 1975 Jun;65(2):588-90 - PubMed
2. Structure. 2009 Jun 10;17(6):789-99 - PubMed
3. Virology. 1966 Jun;29(2):339-45 - PubMed
4. J Mol Biol. 2010 Oct 1;402(4):731-40 - PubMed
5. J Mol Biol. 2007 Aug 10;371(2):374-87 - PubMed
6. BMC Microbiol. 2008 Dec 17;8:227 - PubMed
7. Virology. 1979 Apr 15;94(1):10-23 - PubMed
8. J Biol Chem. 1965 Jan;240:384-90 - PubMed
9. Virology. 2011 Mar 15;411(2):393-415 - PubMed
10. Nature. 2006 Feb 2;439(7076):612-6 - PubMed
11. Structure. 2007 Jul;15(7):807-12 - PubMed
12. Virology. 1973 Mar;52(1):160-73 - PubMed
13. EMBO J. 2005 Jun 15;24(12):2087-95 - PubMed
14. J Virol. 1977 Jul;23(1):91-7 - PubMed
15. Virology. 1973 Mar;52(1):148-59 - PubMed
16. J Mol Biol. 1967 Dec 28;30(3):445-55 - PubMed
17. Vet Res. 2011 Feb 14;42:34 - PubMed
18. Gene. 1991 Dec 1;108(1):1-6 - PubMed
19. Virology. 2007 Dec 20;369(2):234-44 - PubMed
20. Adv Virus Res. 1998;51:135-201 - PubMed
21. Proc Natl Acad Sci U S A. 1967 Feb;57(2):284-91 - PubMed
22. Proc Natl Acad Sci U S A. 2013 Jul 23;110(30):12301-6 - PubMed
23. J Bacteriol. 2009 Jan;191(1):135-40 - PubMed
24. Science. 2006 Jun 23;312(5781):1791-5 - PubMed
25. J Mol Biol. 1997 Apr 11;267(4):865-80 - PubMed
26. Virology. 1958 Feb;5(1):68-91 - PubMed
27. Nature. 2008 Feb 28;451(7182):1130-4 - PubMed
28. J Struct Biol. 2009 Jan;165(1):1-9 - PubMed

Publication Types

• Journal Article

Grant support

• R15 GM052696/GM/NIGMS NIH HHS/United States