Automated C-Terminal Sequencing of Peptides and Proteins

  • Jerome M. Bailey
  • Miro Rusnak
  • John E. Shively


The development of a chemical method for the sequential degradation of a protein or peptide from the carboxy-terminus is a goal of our laboratory. Such a method, in addition to complementing existing N-terminal methods of degradation, would be invaluable for the sequence analysis of proteins with naturally occurring N-terminal blocking groups and for the detection of post-translational processing at the carboxy-terminus of expressed gene products. Although several methods for a sequential C-terminal degradation have been proposed (Ward, 1986; Rangarajan, 1988), the thiocyanate method based on the procedure originally published by Schlack and Kumpf (1926) has been the most widely studied. Recent work in our laboratory, introducing new reagents for the derivatization of the C-terminal amino acid (Hawke et al., 1987) and for the specific cleavage of the derivatized amino acid (Bailey and Shively, 1991; Bailey et al., 1992), has suggested that an automated chemical method for the sequential degradation of polypeptides from the carboxy-terminus, analogous to the Edman method for amino terminal degradation, may be feasible.


Acetic Anhydride Derivatized Amino Acid Amino Acid Ester Continuous Flow Reactor Ethyl Chloroformate 
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  1. Bailey, J. M., Shenoy, N. R., Ronk, M. and Shively, J. E., 1992 Automated carboxy-terminal sequence analysis of peptides, Protein Science 1:68–80.PubMedCrossRefGoogle Scholar
  2. Bailey, J. M. and Shively, J. E., 1990 Carboxy-terminal sequencing: Formation and hydrolysis of C-terminal peptidylthiohydantoins, Biochemistry 29:3145.PubMedCrossRefGoogle Scholar
  3. Bailey, J. M. and Shively, J. E., 1991 Carboxy terminal sequencing: Automation and application to the solid phase, in: Techniques in Protein Chemistry: II, Villafranca, J. J., ed., Academic Press, Inc. New York. pp. 115–129.Google Scholar
  4. Calaycay, J., Rusnak, M. and Shively, J. E., 1991 Microsequencing analysis of peptides and proteins IX, An improved, compact, automated instrument, Anal. Biochem. 192:23.PubMedCrossRefGoogle Scholar
  5. Hawke, D. H., Lahm, H.-W., Shively, J. E. and Todd, C. W., 1987 Microsequence analysis of peptides and proteins: Trimethylsilylisothiocyanate as a reagent for COOH-terminal analysis, Anal. Biochem. 166:298.PubMedCrossRefGoogle Scholar
  6. Hawke, D. H., and Harris, D. C. and Shively, J. E., 1985 Microsequence analysis of peptides and proteins V. Design and performance of a novel gas-liquid phase instrument. Anal. Biochem. 147:315.PubMedCrossRefGoogle Scholar
  7. Husek, P., 1991 Rapid derivatization and gas Chromatographic determination of amino acids, J. Chromatogr. 552:289.CrossRefGoogle Scholar
  8. Kenner, G. W., Khorana, H. G. and Stedman, R. J., 1953 Peptides, Part IV, Selective removal of the C-terminal residue as a thiohydantoin. The use of diphenylphosphoroisothiocyanatidate, Chem. Soc. Jour. (London) 673-678.Google Scholar
  9. Rangarajan, M. 1988 Chemical methods of amino acid sequence analysis from carboxyterminal end, in: Protein/Peptide Sequence Analysis: Current Methodologies, Brown, A. S., ed., CRC Press, Bora Raton, FA., pp. 135–144.Google Scholar
  10. Schlack, P. and Kumpf, W., 1926 Uber eine neue methode zur Ermittlung der konstitution von peptiden, Z. Physiol. Chem. 154:125.CrossRefGoogle Scholar
  11. Shenoy, N. R., Bailey, J. M. and Shively, J. E., 1992 Carboxylic acid modified polyethylene: A novel support for the covalent immobilization of polypeptides for C-terminal sequencing, Protein Science 1:58.PubMedCrossRefGoogle Scholar
  12. Shively, J. E., Miller, P. and Ronk, M., 1987 Microsequence analysis of peptides and proteins VI. A continuous flow reactor for sample concentration and sequence analysis, Anal. Biochem. 163:517.PubMedCrossRefGoogle Scholar
  13. Ward, C. W., 1986 Carboxyl terminal sequence analysis, in: Practical Protein Chemistry — A Handbook, Darbre, A., ed., John Wiley and Sons, Ltd, Chichester, New York, pp. 491–525.Google Scholar

Copyright information

© Springer Science+Business Media New York 1993

Authors and Affiliations

  • Jerome M. Bailey
    • 1
  • Miro Rusnak
    • 1
  • John E. Shively
    • 1
  1. 1.Division of ImmunologyBeckman Research Institute of the City of HopeDuarteUSA

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