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. 1978 Jul;27(1):218–226. doi: 10.1128/jvi.27.1.218-226.1978

Measurements of the molecular size of the simian virus 40 large T antigen.

J D Griffin, S Light, D M Livingston
PMCID: PMC354154  PMID: 211251

Abstract

A measure of the molecular weight of the large simian virus 40 T antigen was sought by SDS-polyacrylamide gel electrophoresis, random-coil chromatography, and sedimentation-velocity analysis in a density gradient. Large T antigen obtained from a simian virus 40-transformed human cell line either by immunoprecipitation or by standard preparatory methods migrated like a 94,000-molecular-weight (approximately 94K) polypeptide in SDS-gels but was found to have an approximate was observed with T antigen obtained from lytically infected monkey cells. In view of the strong theoretical basis for the guanidine method and the agreement with the sedimentation data, these findings suggest that the molecular weight of this protein is approximately 75 to 80K as opposed to 94 to 100K and, therefore, that considerably less than the entire early region of simian virus 40 is required to encode it. This size estimate is in keeping with earlier results which revealed a normal-size T antigen in cells infected with viable deletion mutants lacking as much as 10% of the early region.

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Selected References

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  1. Ahmad-Zadeh C., Allet B., Greenblatt J., Weil R. Two forms of simian-virus-40-specific T-antigen in abortive and lytic infection. Proc Natl Acad Sci U S A. 1976 Apr;73(4):1097–1101. doi: 10.1073/pnas.73.4.1097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aloni Y., Dhar R., Laub O., Horowitz M., Khoury G. Novel mechanism for RNA maturation: the leader sequences of simian virus 40 mRNA are not transcribed adjacent to the coding sequences. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3686–3690. doi: 10.1073/pnas.74.9.3686. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berget S. M., Moore C., Sharp P. A. Spliced segments at the 5' terminus of adenovirus 2 late mRNA. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3171–3175. doi: 10.1073/pnas.74.8.3171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  5. Brugge J. S., Butel J. S. Role of simian virus 40 gene A function in maintenance of transformation. J Virol. 1975 Mar;15(3):619–635. doi: 10.1128/jvi.15.3.619-635.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. CRAVEN G. R., STEERS E., Jr, ANFINSEN C. B. PURIFICATION, COMPOSITION, AND MOLECULAR WEIGHT OF THE BETA-GALACTOSIDASE OF ESCHERICHIA COLI K12. J Biol Chem. 1965 Jun;240:2468–2477. [PubMed] [Google Scholar]
  7. Carroll R. B., Smith A. E. Monomer molecular weight of T antigen from simian virus 40-infected and transformed cells. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2254–2258. doi: 10.1073/pnas.73.7.2254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chou J. Y., Martin R. G. Complementation analysis of simian virus 40 mutants. J Virol. 1974 May;13(5):1101–1109. doi: 10.1128/jvi.13.5.1101-1109.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chou J. Y., Martin R. G. DNA infectivity and the induction of host DNA synthesis with temperature-sensitive mutants of simian virus 40. J Virol. 1975 Jan;15(1):145–150. doi: 10.1128/jvi.15.1.145-150.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chow L. T., Gelinas R. E., Broker T. R., Roberts R. J. An amazing sequence arrangement at the 5' ends of adenovirus 2 messenger RNA. Cell. 1977 Sep;12(1):1–8. doi: 10.1016/0092-8674(77)90180-5. [DOI] [PubMed] [Google Scholar]
  11. Crawford L. V., Cole C. N., Smith A. E., Paucha E., Tegtmeyer P., Rundell K., Berg P. Organization and expression of early genes of simian virus 40. Proc Natl Acad Sci U S A. 1978 Jan;75(1):117–121. doi: 10.1073/pnas.75.1.117. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Del Villano B. C., Defendi V. Characterization of the SV40 T antigen. Virology. 1973 Jan;51(1):34–46. doi: 10.1016/0042-6822(73)90363-2. [DOI] [PubMed] [Google Scholar]
  13. Dunn A. R., Hassell J. A. A novel method to map transcripts: evidence for homology between an adenovirus mRNA and discrete multiple regions of the viral genome. Cell. 1977 Sep;12(1):23–36. doi: 10.1016/0092-8674(77)90182-9. [DOI] [PubMed] [Google Scholar]
  14. Eckhart W. Complementation and transformation by temperature-sensitive mutants of polyoma virus. Virology. 1969 May;38(1):120–125. doi: 10.1016/0042-6822(69)90133-0. [DOI] [PubMed] [Google Scholar]
  15. Feunteun J., Sompayrac L., Fluck M., Benjamin T. Localization of gene functions in polyoma virus DNA. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4169–4173. doi: 10.1073/pnas.73.11.4169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Fish W. W., Mann K. G., Tanford C. The estimation of polypeptide chain molecular weights by gel filtration in 6 M guanidine hydrochloride. J Biol Chem. 1969 Sep 25;244(18):4989–4994. [PubMed] [Google Scholar]
  17. Fluck M. M., Staneloni R. J., Benjamin T. L. Hr-t and ts-a: two early gene functions of polyoma virus. Virology. 1977 Apr;77(2):610–624. doi: 10.1016/0042-6822(77)90486-x. [DOI] [PubMed] [Google Scholar]
  18. GILDEN R. V., CARP R. I., TAGUCHI F., DEFEND V. THE NATURE AND LOCALIZATION OF THE SV 40-INDUCED COMPLEMENT-FIXING ANTIGEN. Proc Natl Acad Sci U S A. 1965 Mar;53:684–692. doi: 10.1073/pnas.53.3.684. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Greenblatt J. F., Allet B., Weil R., Ahmad-Zadeh C. Synthesis of the tumour antigen and the major capsid protein of simian virus 40 in a cell-free system derived from Escherichia coli. J Mol Biol. 1976 Dec;108(2):361–379. doi: 10.1016/s0022-2836(76)80125-8. [DOI] [PubMed] [Google Scholar]
  21. Henderson I. C., Livingston D. M. Partial purification and characterization of the SV40 T antigen. Cell. 1974 Sep;3(1):65–70. doi: 10.1016/0092-8674(74)90041-5. [DOI] [PubMed] [Google Scholar]
  22. Ito Y., Brocklehurst J. R., Dulbecco R. Virus-specific proteins in the plasma membrane of cells lytically infected or transformed by pol-oma virus. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4666–4670. doi: 10.1073/pnas.74.10.4666. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jessel D., Hudson J., Landau T., Tenen D., Livingston D. M. Interaction of partially purified simian virus 40 T antigen with circular viral DNA molecules. Proc Natl Acad Sci U S A. 1975 May;72(5):1960–1964. doi: 10.1073/pnas.72.5.1960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Khoury G., Howley P., Nathans D., Martin M. Posttranscriptional selection of simian virus 40-specific RNA. J Virol. 1975 Feb;15(2):433–437. doi: 10.1128/jvi.15.2.433-437.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Khoury G., May E. Regulation of early and late simian virus 40 transcription: overproduction of early viral RNA in the absence of a functional T-antigen. J Virol. 1977 Jul;23(1):167–176. doi: 10.1128/jvi.23.1.167-176.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Klessig D. F. Two adenovirus mRNAs have a common 5' terminal leader sequence encoded at least 10 kb upstream from their main coding regions. Cell. 1977 Sep;12(1):9–21. doi: 10.1016/0092-8674(77)90181-7. [DOI] [PubMed] [Google Scholar]
  27. Kuchino T., Yamaguchi N. Characterization of T antigen in cells infected with a temperature-sensitive mutant of simian virus 40. J Virol. 1975 Jun;15(6):1302–1307. doi: 10.1128/jvi.15.6.1302-1307.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  29. Lewis J. B., Anderson C. W., Atkins J. F. Further mapping of late adenovirus genes by cell-free translation of RNA selected by hybridization to specific DNA fragments. Cell. 1977 Sep;12(1):37–44. doi: 10.1016/0092-8674(77)90183-0. [DOI] [PubMed] [Google Scholar]
  30. MAEHLY A. C., CHANCE B. The assay of catalases and peroxidases. Methods Biochem Anal. 1954;1:357–424. doi: 10.1002/9780470110171.ch14. [DOI] [PubMed] [Google Scholar]
  31. MARTIN R. G., AMES B. N. A method for determining the sedimentation behavior of enzymes: application to protein mixtures. J Biol Chem. 1961 May;236:1372–1379. [PubMed] [Google Scholar]
  32. Martin R. G., Chou J. Y. Simian virus 40 functions required for the establishment and maintenance of malignant transformation. J Virol. 1975 Mar;15(3):599–612. doi: 10.1128/jvi.15.3.599-612.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Martin R. G., Oppenheim A. Initiation points for DNA replication in nontransformed and simian virus 40-transformed Chinese hamster lung cells. Cell. 1977 Aug;11(4):859–869. doi: 10.1016/0092-8674(77)90297-5. [DOI] [PubMed] [Google Scholar]
  34. Osborn M., Weber K. Simian virus 40 gene A function and maintenance of transformation. J Virol. 1975 Mar;15(3):636–644. doi: 10.1128/jvi.15.3.636-644.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Panyim S., Chalkley R. The molecular weights of vertebrate histones exploiting a modified sodium dodecyl sulfate electrophoretic method. J Biol Chem. 1971 Dec 25;246(24):7557–7560. [PubMed] [Google Scholar]
  36. Potter C. W., McLaughlin B. C., Oxford J. S. Simian virus 40-induced T and tumor antigens. J Virol. 1969 Nov;4(5):574–579. doi: 10.1128/jvi.4.5.574-579.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Prives C., Gilboa E., Revel M., Winocour E. Cell-free translation of simian virus 40 early messenger RNA coding for viral T-antigen. Proc Natl Acad Sci U S A. 1977 Feb;74(2):457–461. doi: 10.1073/pnas.74.2.457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Reed S. I., Stark G. R., Alwine J. C. Autoregulation of simian virus 40 gene A by T antigen. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3083–3087. doi: 10.1073/pnas.73.9.3083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Rundell K., Collins J. K., Tegtmeyer P., Ozer H. L., Lai C. J., Nathans D. Identification of simian virus 40 protein A. J Virol. 1977 Feb;21(2):636–646. doi: 10.1128/jvi.21.2.636-646.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Shenk T. E., Carbon J., Berg P. Construction and analysis of viable deletion mutants of simian virus 40. J Virol. 1976 May;18(2):664–671. doi: 10.1128/jvi.18.2.664-671.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sompayrac L., Danna K. J. Measurements of the genome sizes of simian virus 40 and polyoma virus. J Virol. 1977 Nov;24(2):695–700. doi: 10.1128/jvi.24.2.695-700.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Staneloni R. J., Fluck M. M., Benjamin T. L. Host range selection of transformation-defective hr-t mutants of polyoma virus. Virology. 1977 Apr;77(2):598–609. doi: 10.1016/0042-6822(77)90485-8. [DOI] [PubMed] [Google Scholar]
  43. Subramanian K. N., Dhar R., Weissman S. M. Nucleotide sequence of a fragment of SV40 DNA that contains the origin of DNA replication and specifies the 5' ends of "early" and "late" viral RNA. III. Construction of the total sequence of EcoRII-G fragment of SV40 DNA. J Biol Chem. 1977 Jan 10;252(1):355–367. [PubMed] [Google Scholar]
  44. Tegtmeyer P. Function of simian virus 40 gene A in transforming infection. J Virol. 1975 Mar;15(3):613–618. doi: 10.1128/jvi.15.3.613-618.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tegtmeyer P., Rundell K., Collins J. K. Modification of simian virus 40 protein A. J Virol. 1977 Feb;21(2):647–657. doi: 10.1128/jvi.21.2.647-657.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Tegtmeyer P., Schwartz M., Collins J. K., Rundell K. Regulation of tumor antigen synthesis by simain virus 40 gene A. J Virol. 1975 Jul;16(1):168–178. doi: 10.1128/jvi.16.1.168-178.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Tegtmeyer P. Simian virus 40 deoxyribonucleic acid synthesis: the viral replicon. J Virol. 1972 Oct;10(4):591–598. doi: 10.1128/jvi.10.4.591-598.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Tenen D. G., Baygell P., Livingston D. M. Thermolabile T (tumor) antigen from cells transformed by a temperature-sensitive mutant of simian virus 40. Proc Natl Acad Sci U S A. 1975 Nov;72(11):4351–4355. doi: 10.1073/pnas.72.11.4351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Tenen D. G., Garewal H., Haines L. L., Hudson J., Woodard V., Light S., Livingston D. M. Purification of simian virus 40 tumor antigen from a line of simian virus 40-transformed human cells. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3745–3749. doi: 10.1073/pnas.74.9.3745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Tenen D. G., Martin R. G., Anderson J., Livingston D. M. Biological and biochemical studies of cells transformed by simian virus 40 temperature-sensitive gene A mutants and A mutant revertants. J Virol. 1977 Apr;22(1):210–218. doi: 10.1128/jvi.22.1.210-218.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Tung J. S., Knight C. A. Effect of charge on the determination of molecular weight of proteins by gel electrophoresis in SDS. Biochem Biophys Res Commun. 1971 Mar 19;42(6):1117–1121. doi: 10.1016/0006-291x(71)90020-9. [DOI] [PubMed] [Google Scholar]

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