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Target Size Analysis of Cell Surface Receptors and Adenylate Cyclase

  • Werner Schlegel
  • Ellis S. Kempner
Part of the Methodological Surveys in Biochemistry and Analysis book series (MSBA, volume 13)

Abstract

Radiation inactivation with target size analysis [1] allows determination of the molecular size of enzymes or receptors with no need for purification or solubilization. Known doses of high-energy radiation, viz. electrons from a linear accelerator, are delivered homogeneously to a sample of a crude enzyme or receptor preparation. To avoid indirect effects of radiation the sample has to be either dry (lyophilized) or kept frozen at a constant temperature. From the loss of function — catalytic activity or specific binding — as a function of radiation dose, the target size of an enzyme or a receptor can be evaluated.

A literature review [2] showed that this method measures either the size of subunits or subunit assemblies or the complete size of the enzyme molecule. We proposed that the target size corresponds to the size of the functional unit, i.e. the minimal assembly of protein subunits required for a given enzymatic or specific binding function [2]. This concept has been verified by target size analysis of soluble polyenzyme clusters [3]. It can be shown that within the same cluster, target sizes for distinct functions are different, indicating which subunits or subunit assemblies constitute the functional unit for a given catalytic activity. This article concerns technical problems, the theoretical aspects, and examples for adenylate cyclase in liver/ fat cells/turkey erythrocyte membranes etc. to illustrate implications of the functional unit concept for interpreting target data, and how radiation inactivation complements biochemical approaches.

Keywords

Adenylate Cyclase Functional Unit Primary Ionization Target Size Target Analysis 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. 1.
    Lea, D.E. (1955) Actions of Radiations on Living Cells, 2nd edn., Cambridge University Press, Cambridge, 416 pp.Google Scholar
  2. 2.
    Kempner, E.S. & Schlegel, W. (1979) Anal. Biochem. 92, 2–10.CrossRefGoogle Scholar
  3. 3.
    Kempner, E.S., Miller, J.H., Schlegel, W. & Hearon, J.Z. (1980) J. Biol. Chem. 255, 6826–6831.Google Scholar
  4. 4.
    Brodsky, A. (1978) CRC Handbook of Radiation Measurement and Protection, Vol. 1, Physical Science and Engineering Data, CRC Press, W. Palm Beach, Florida, 693 pp.Google Scholar
  5. 5.
    Nielsen, T.B., Lad, P.M., Preston, M.S., Kempner, E.S., Schlegel, W.& Rodbell, M. (198) Proc. Nat. Acad. Sci. 78, 722–726.Google Scholar
  6. 6.
    Kepner, G.R. & Macey, R.I. (1968) Biochim. Biophys. Acta 163, 188–203.CrossRefGoogle Scholar
  7. 7.
    Parkinson, D. & Callingham, B.A. (1982) Rad. Res. 90, 252–259.CrossRefGoogle Scholar
  8. 8.
    Harmon, J.T., Kahn, C.R., Kempner, E.S. & Schlegel, W. (1980) J. Biol. Chem. 255, 3412–3419.Google Scholar
  9. 9.
    Harmon, J.T., Kempner, E.S. & Kahn, C.R. (1980) J. Biol. Chem. 256, 7719–7722.Google Scholar
  10. 10.
    Doble, A. & Iversen, L.L. (1982) Nature 295, 522–523.CrossRefGoogle Scholar
  11. 11.
    Fewtrell, C., Kempner, E., Poy, G. & Metzger, H. (1981) Biochemistry 20, 6589–6594.CrossRefGoogle Scholar
  12. 12.
    Pollet, R.J., Kempner, E.S., Standaert, M.L. & Haase, B.A. (1982) J. Biol. Chem. 257, 894–898.Google Scholar
  13. 13.
    Schlegel, W., Kempner, E.S. & Rodbell, M. (1979) J. Biol. Chem. 254, 5168–5176.Google Scholar
  14. 14.
    Innerarity, T.L., Kempner, E.S., Hui, D.Y. & Mahley, R.H. (1981) Proc. Nat. Acad. Sci. 78, 4378–4382.CrossRefGoogle Scholar
  15. 15.
    Steer, C.J., Kempner, E.S. & Ashwell, G. (1981) J. Biol. Chem. 256, 5851–5856.Google Scholar
  16. 16.
    Paul, S.M., Kempner, E.S. & Skolnick, P. (1981) Eur. J. Pharmacol, 76, 465–466.CrossRefGoogle Scholar
  17. 17.
    Chang, L., Barnard, E.A., Lo, M.M.S. & Dolly, J.O. (1981) FEBS Lett. 126, 309–312.CrossRefGoogle Scholar
  18. 18.
    Barnard, E.A., Dolly, J.O., Lo, M. & Mantle, T. (1978) Biochem. Soc. Trans. 6, 649–651.Google Scholar
  19. 19.
    Lo, M.M.S., Barnard, E.A. & Dolly, J.O. (1982) Biochemistry 21, 2210–2217.CrossRefGoogle Scholar
  20. 20.
    Levinson, S.R. & Ellory, J.C (1973) Nature 245, 122–123.CrossRefGoogle Scholar
  21. 21.
    Rodbell, M. (1980) Nature 284, 17–22.CrossRefGoogle Scholar
  22. 22.
    Houslay, M.D., Ellory, J.C., Smith, G.A., Hesketh, T.R., Stein, J.M., Warren, J.B. & Metcalfe, J.C. (1977) Biochim. Biophys. Acta 467, 208–219.CrossRefGoogle Scholar
  23. 23.
    Martin, B.R., Stein, J.M., Kennedy, E.L., Doberska, C.A. & Metcalfe, J.C (1979) Biochem. J. 184, 253–260.Google Scholar
  24. 24.
    Martin, B.R., Stein, J.M., Kennedy, E.L. & Doberska, C.A. (1980) Biochem. J. 188, 137–140.Google Scholar
  25. 25.
    Schlegel, W., Cooper, D.M.F. & Rodbell, M. (1980) Arch. Biochem. Biophys. 201, 678–682.CrossRefGoogle Scholar
  26. 26.
    Cassel, D. & Selinger, Z. (1978) Proc. Nat. Acad. Sci. 75, 4155–4159.CrossRefGoogle Scholar
  27. 27.
    Iyengar, R. & Birnbaumer, L. (1981) J. Biol. Chem. 256, 11036–11041.Google Scholar

Copyright information

© Plenum Press, New York 1984

Authors and Affiliations

  • Werner Schlegel
    • 1
  • Ellis S. Kempner
    • 2
  1. 1.Department of MedicineFondation pour Recherches MedicalesGenève 4Switzerland
  2. 2.National Institutes of HealthBethesdaUSA

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