Correlated Measurement of Endogenous ATE1 Activity on Native Acceptor Proteins in Tissues and Cultured Cells to Detect Cellular Aging

  • Hideko KajiEmail author
  • Akira KajiEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1337)


Following our early discovery of arginylation in 1963, we have performed several studies to correlate its activity with essential biological processes. We employed cell- and tissue-based assays to detect both the level of acceptor proteins and the level of ATE1 activity under different conditions. Remarkably, in these assays, we found a close correlation between arginylation and aging, a discovery that we believe has longer-term implications in uncovering the importance of ATE1 in normal biology and disease therapies. Here we describe the original methods we used to measure ATE1 activity in tissues and correlate it with key biological events.

Key words

Arginylation Aging Cell proliferation SV40 transformation ATE1 Arg acceptor proteins 


  1. 1.
    Kaji A, Kaji H, Novelli GD (1963) A soluble amino acid incorporating system. Biochem Biophys Res Commun 10:406–409CrossRefPubMedGoogle Scholar
  2. 2.
    Kaji A, Kaji H, Novelli GD (1965) Soluble amino acid-incorporation system. I. Preparation of the system and the nature of the reaction. J Biol Chem 240:1185–1191PubMedGoogle Scholar
  3. 3.
    Kaji A, Kaji H, Novelli GD (1965) Soluble amino acid-incorporating system. II. Soluble nature of the system and the characterization of the radioactive product. J Biol Chem 240:1192–1197PubMedGoogle Scholar
  4. 4.
    Kaji H, Novelli GD, Kaji A (1963) A soluble amino acid-incorporating system from rat liver. Biochim Biophys Acta 76:474–477CrossRefPubMedGoogle Scholar
  5. 5.
    Momose K, Kaji A (1966) Soluble amino acid incorporating-system. III. Further studies on the product and its relation to the ribosomal system for incorporation. J Biol Chem 241:3294–3307PubMedGoogle Scholar
  6. 6.
    Tanaka Y, Kaji H (1974) Incorporation of arginine by soluble extracts of ascites tumor cells and regenerating rat liver. Cancer Res 34(9):2204–2208PubMedGoogle Scholar
  7. 7.
    Rao P, Kaji H (1977) Comparative studies on isoaccepting arginyl tRNAs from transformed cells and their utilization in post-translational protein modification. Arch Biochem Biophys 181(2):591–595CrossRefPubMedGoogle Scholar
  8. 8.
    Rao P, Kaji H (1977) Effect of temperature on arginine incorporation by ribosomeless extracts of cells transformed by a temperature-sensitive mutant of Rous sarcoma virus. Biochim Biophys Acta 477(4):394–403CrossRefPubMedGoogle Scholar
  9. 9.
    Gabius HJ, Engelhardt R, Deerberg F, Cramer F (1983) Age-related changes in different steps of protein synthesis of liver and kidney of rats. FEBS Lett 160(1-2):115–118CrossRefPubMedGoogle Scholar
  10. 10.
    Bongiovanni G, Fissolo S, Barra HS, Hallak ME (1999) Posttranslational arginylation of soluble rat brain proteins after whole body hyperthermia. J Neurosci Res 56(1):85–92CrossRefPubMedGoogle Scholar
  11. 11.
    Hayflick L (1965) The limited in vitro lifetime of human diploid cell strains. Exp Cell Res 37:614–636CrossRefPubMedGoogle Scholar
  12. 12.
    Shein HM, Enders JF (1962) Multiplication and cytopathogenicity of Simian vacuolating virus 40 in cultures of human tissues. Proc Soc Exp Biol Med 109:495–500CrossRefPubMedGoogle Scholar
  13. 13.
    Koprowski H, Croce CM (1977) Tumorigenicity of simian virus 40-transformed human cells and mouse–human hybrids in nude mice. Proc Natl Acad Sci U S A 74(3):1142–1146PubMedCentralCrossRefPubMedGoogle Scholar
  14. 14.
    Kaji H, Hara H, Lamon KD (1980) Fixation of cellular aging processes by SV40 virus transformation. Mech Ageing Dev 12(2):197–209CrossRefPubMedGoogle Scholar
  15. 15.
    Kaji H, Rao P (1976) Membrane modification by arginyl tRNA. FEBS Lett 66(2):194–197CrossRefPubMedGoogle Scholar
  16. 16.
    Bongiovanni G, Fidelio GD, Barra HS, Hallak ME (1995) The post-translational incorporation of arginine into a beta-amyloid peptide increases the probability of alpha-helix formation. Neuroreport 7(1):326–328PubMedGoogle Scholar
  17. 17.
    Hallak ME, Bongiovanni G, Barra HS (1991) The posttranslational arginylation of proteins in different regions of the rat brain. J Neurochem 57(5):1735–1739CrossRefPubMedGoogle Scholar
  18. 18.
    Brower CS, Piatkov KI, Varshavsky A (2013) Neurodegeneration-associated protein fragments as short-lived substrates of the N-end rule pathway. Mol Cell 50(2):161–171PubMedCentralCrossRefPubMedGoogle Scholar
  19. 19.
    Lamon KD, Chiger JL, Kaji H (1982) Effects of hydrocortisone and aspirin on protein synthesis and post-translational protein modification in cultured cells. Biochem Pharmacol 31(11):2047–2052CrossRefPubMedGoogle Scholar
  20. 20.
    Lamon KD, Kaji H (1980) Arginyl-tRNA transferase activity as a marker of cellular aging in peripheral rat tissues. Exp Gerontol 15(1):53–64CrossRefPubMedGoogle Scholar
  21. 21.
    Lamon KD, Vogel WH, Kaji H (1980) Stress-induced increases in rat brain arginyl-tRNA transferase activity. Brain Res 190(1):285–287CrossRefPubMedGoogle Scholar
  22. 22.
    Saha S, Mundia MM, Zhang F, Demers RW, Korobova F, Svitkina T, Perieteanu AA, Dawson JF, Kashina A (2010) Arginylation regulates intracellular actin polymer level by modulating actin properties and binding of capping and severing proteins. Mol Biol Cell 21(8):1350–1361PubMedCentralCrossRefPubMedGoogle Scholar
  23. 23.
    Hallak ME, Bongiovanni G (1997) Posttranslational arginylation of brain proteins. Neurochem Res 22(4):467–473CrossRefPubMedGoogle Scholar
  24. 24.
    Saha S, Kashina A (2011) Posttranslational arginylation as a global biological regulator. Dev Biol 358:1–8PubMedCentralCrossRefPubMedGoogle Scholar
  25. 25.
    Soffer RL (1970) Enzymatic modification of proteins. II. Purification and properties of the arginyl transfer ribonucleic acid-protein transferase from rabbit liver cytoplasm. J Biol Chem 245(4):731–737PubMedGoogle Scholar
  26. 26.
    Mans RJ, Novelli GD (1961) Measurement of the incorporation of radioactive amino acids into protein by a filter-paper disk method. Arch Biochem Biophys 94:48–53CrossRefGoogle Scholar
  27. 27.
    Tegtmeyer P (1975) Function of simian virus 40 gene A in transforming infection. J Virol 15(3):613–618PubMedCentralPubMedGoogle Scholar
  28. 28.
    Girardi AJ, Weinstein D, Moorhead PS (1966) SV40 transformation of human diploid cells. A parallel study of viral and karyologic parameters. Ann Med Exp Biol Fenn 44(2):242–254PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Biochemistry and Molecular Biology, Jefferson Medical CollegeThomas Jefferson UniversityPhiladelphiaUSA
  2. 2.Department of Microbiology, Perelman School of MedicineUniversity of PennsylvaniaPhiladelphiaUSA

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