Abstract
Thymidine kinase enzymes isolated from a variety of sources are generally considered to have a native molecular weight of 80–90 kDa composed of two 40–45 kDa subunits. Although these parameters may accurately describe the atypical deoxypyrimidine kinases expressed by members of the Herpesviridae, the nucleotide sequences of thymidine kinase genes isolated from human, mouse, chicken and a variety of poxviruses (vaccinia virus, monkeypox virus, variola virus, fowlpox virus and capripoxvirus) predict molecular weights on the order of 20–25 kDa for the derived primary translation products. To resolve this apparent dilemma, velocity sedimentation centrifugation, gel filtration chromatography and protein cross-linking procedures were employed to provide experimental evidence that enzymatically-active vaccinia virus thymidine kinase is a homotetrameric complex of 20 kDa monomers with a native Mr of 80 kDa.
Abbreviations: VV, Vaccinia Virus; TK, thymidine kinase; HSV, Herpes Simplex Virus; SDS, sodium dodecyl sulfate; TCA, trichloroacetic acid
References
- 1.Flemington E., Bradshaw H.D., Jr., Traina-Dorge V., Slagel V., Deininger P.L. Gene. 1987;52:267–277. doi: 10.1016/0378-1119(87)90053-9. [DOI] [PubMed] [Google Scholar]
- 2.Lin P.F., Lieberman H.B., Yeh D.B., Xu T., Zhao S.Y., Ruddle F.H. Mol. Cell. Biol. 1985;5:3149–3156. doi: 10.1128/mcb.5.11.3149. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Kwoh T.J., Engler J.A. Nucl. Acids Res. 1984;12:3959–3971. doi: 10.1093/nar/12.9.3959. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Nunberg J.H., Wright D.K., Cole G.E., Petrovskis E.A., Post L.E., Compton T., Gilbert J.H. J. Virol. 1989;63:3240–3249. doi: 10.1128/jvi.63.8.3240-3249.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Valerie K., Stevens J., Lynch M., Henderson E.E., de Riel J.K. Nucl. Acids. Res. 1986;14:8637–8654. doi: 10.1093/nar/14.21.8637. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Weir J.P., Moss B. J. Virol. 1983;46:530–537. doi: 10.1128/jvi.46.2.530-537.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Esposito J.J., Knight J.C. Virology. 1984;135:561–567. doi: 10.1016/0042-6822(84)90212-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Boyle D.B., Coupar B.E.H., Gibbs A.J., Seigman L.J., Both G.W. Virology. 1987;156:355–365. doi: 10.1016/0042-6822(87)90415-6. [DOI] [PubMed] [Google Scholar]
- 9.Gershon P.D., Black D.N. J. Gen. Virol. 1989;70:525–533. doi: 10.1099/0022-1317-70-3-525. [DOI] [PubMed] [Google Scholar]
- 10.Traut T.W. CRC Crit. Rev. Biochem. 1988;23:121–169. doi: 10.3109/10409238809088318. [DOI] [PubMed] [Google Scholar]
- 11.Hruby D.E., Ball L.A. J. Virol. 1982;43:403–409. doi: 10.1128/jvi.43.2.403-409.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Wilson E.M., Franke C.A., Black M.E., Hruby D.E. Gene. 1989;77:69–78. doi: 10.1016/0378-1119(89)90360-0. [DOI] [PubMed] [Google Scholar]
- 13.Hruby D.E., Ball L.A. Virology. 1981;113:594–601. doi: 10.1016/0042-6822(81)90187-2. [DOI] [PubMed] [Google Scholar]
- 14.Studier F.W. J. Mol. Biol. 1973;79:237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
- 15.Wagner M.J., Sharp J.A., Summers W.C. 3rd ed. Vol. 78. 1981. pp. 1441–1445. (Proc. Natl. Acad. Sci., U.S.A). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Littler E., Arrand J.R. J. Virol. 1988;62:3892–3895. doi: 10.1128/jvi.62.10.3892-3895.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Otsuka H., Kit S. Virology. 1984;135:316–330. doi: 10.1016/0042-6822(84)90189-2. [DOI] [PubMed] [Google Scholar]
- 18.Davison A.J., Scott J.E. J. Gen. Virol. 1986;67:1759–1816. doi: 10.1099/0022-1317-67-9-1759. [DOI] [PubMed] [Google Scholar]
- 19.Kit S., Jorgensen G.N., Liav A., Zaslavsky V. Virology. 1977;77:661–676. doi: 10.1016/0042-6822(77)90490-1. [DOI] [PubMed] [Google Scholar]
- 20.Hruby D.E. Virus Research. 1985;2:151–156. doi: 10.1016/0168-1702(85)90245-x. [DOI] [PubMed] [Google Scholar]
- 21.Sherely J.L., Kelly T.J. J. Biol. Chem. 1988;263:375–382. [PubMed] [Google Scholar]