Skip to main content

HPLC in the Analysis of Peptide Metabolism

  • Protocol

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 251))

Abstract

Bioactive peptides are generally synthesized within large precursor molecules, from which the active moiety must be enzymatically excised by one or more specific peptidases (1). For most peptides, this processing occurs intracellularly, within the secretory pathway, but cosecretory or extracellular processing can also occur. Peptidases also play a critical role in the termination of peptide signals, via cleavage to inactive fragments (2,3). The characterization of the peptidases involved in the generation and metabolism of peptides is thus of critical importance to the understanding of the physiology of specific peptide hormones and neurotransmitters.

This is a preview of subscription content, log in via an institution.

Buying options

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   169.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Docherty, K. and Steiner, D. F. (1982) Post-translational proteolysis in polypeptide hormone biosynthesis. Ann. Rev. Physiol. 44, 625–638.

    Article  CAS  Google Scholar 

  2. Turner, A. J. and Barnes, K. (1994) Neuropeptidases: candidate enzymes and techniques for study. Biochem. Soc. Trans. 22, 122–127.

    PubMed  CAS  Google Scholar 

  3. Schwartz, J.-C, Malfroy, B., and de la Baume, S. (1981) Biological inactivation of enkephalins and the role of enkephalin-dipeptidyl-carboxypeptidase (“enkephalinase”) as neuropeptidase. Life Sci. 29, 1715–1740.

    Article  PubMed  CAS  Google Scholar 

  4. Chu, T. G. and Orlowski, M. (1984) Active site directed N-carboxymethyl peptide inhibitors of a soluble metalloendopeptidase from rat brain. Biochemistry 23, 3598–3603.

    Article  PubMed  CAS  Google Scholar 

  5. Shrimpton, C. N., Abbenante, G., Lew, R. A., and Smith, A. I. (2000) Development and characterisation of novel potent and stable inhibitors of endopeptidase EC 34.24.15. Biochem. J. 345, 351–356.

    Article  PubMed  CAS  Google Scholar 

  6. Vincent, B., Jiracek, J., Noble, F., et al. (1997) Effect of a novel selective and potent phosphinic peptide inhibitor of endopeptidase 34.24.16 on neurotensin-induced analgesia and neuronal inactivation. Brit. J. Pharmacol. 121, 705–710.

    Article  CAS  Google Scholar 

  7. Jiracek, J., Yiotakis, A., Vincent, B., et al. (1995) Development of highly potent and selective phosphinic peptide inhibitors of zinc endopeptidase 24.15 using combinatorial chemistry. J. Biol. Chem. 270, 21,701–21,706.

    Article  PubMed  CAS  Google Scholar 

  8. Dauch, P., Vincent, J.-P., and Checler, F. (1991) Specific inhibition of endopeptidase 24.16 by dipeptides. Eur. J. Biochem. 202, 269–276.

    Article  PubMed  CAS  Google Scholar 

  9. Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J. (1951) Protein measurement surement with the folin phenol reagent. J. Biol. Chem. 193, 265–275.

    PubMed  CAS  Google Scholar 

  10. Skidgel, R. A. (1992) Bradykinin-degrading enzymes: structure, function, distribution, and potential roles in cardiovascular pharmacology. J. Cardiovasc. Pharmacol. 20 (Suppl. 9), S4–S9.

    Article  PubMed  CAS  Google Scholar 

  11. Hoang, M. V. and Turner, A. J. (1997) Novel activity of endothelin-converting enzyme: hydrolysis of bradykinin. Biochem. J. 327, 23–26.

    PubMed  CAS  Google Scholar 

  12. Smith, A. I. and Lew, R. A. (1995) The combination of high performance liquid chromatography and radioimmunoassay for the characterization of peptide processing pathways, in Peptidases and Neuropeptide Processing (Smith, A. I., ed.), Academic, San Diego, CA, pp. 125–139.

    Chapter  Google Scholar 

  13. Campbell, D. J., Kladis, A., and Duncan, A.-M. (1993) Nephrectomy, converting enzyme inhibition and angiotensin peptides. Hypertension 22, 513–522.

    PubMed  CAS  Google Scholar 

  14. Campbell, D. J., Kladis, A., and Duncan, A.-M. (1993) Bradykinin peptides in kidney, blood, and other tissues of the rat. Hypertension 21, 155–165.

    PubMed  CAS  Google Scholar 

  15. Lawrence, A. C., Evin, G., Kladis, A., and Campbell, D. J. (1990) An alternative strategy for the radioimmunoassay of angiotensin peptides using amino-terminal-directed antisera: measurement of eight angiotensin peptides in human plasma. . J. Hypertension 8, 715–724.

    Article  CAS  Google Scholar 

  16. Dean, R., Murone, C., Lew, R. A., et al. (1997) Localization of bradykinin B2 binding sites in rat kidney following chronic ACE inhibitor treatment. Kidney Int. 52, 1261–1270.

    Article  PubMed  CAS  Google Scholar 

  17. Lew, R. A., Tomoda, F., Evans, R. G., et al. (1996) Synthetic inhibitors of endopeptidase EC 3.4.24.15: potency and stability in vitro and in vivo. Brit. J. Pharmacol. 118, 1269–1277.

    CAS  Google Scholar 

  18. Smith, A. I., Lew, R. A., Shrimpton, C. N., Evans, R. G., and Abbenante, G. (2000) A novel stable inhibitor of endopeptidases EC 3.4.24.15 and 3.4.24.16 potentiates bradykinin-induced hypotension. Hypertension 35, 626–630.

    PubMed  CAS  Google Scholar 

  19. Telford, S. E., Smith, A. I., Lew, R. A., Perich, R. B., Madden, A. C, and Evans, R. G. (1995) Role of angiotensin converting enzyme in the vascular effects of an endopeptidase 24.15 inhibitor. Br. J. Pharmacol. 114, 1185–1192.

    PubMed  CAS  Google Scholar 

  20. Moeller I., Lew, R. A., Albiston, A. L., Mendelsohn, F A. O., and Chai, S.-Y. (1999) A globin fragment, LVV-hemorphin-7, induces [3H]thymidine incorporation in a neuronal cell line via the AT4 receptor. J. Neurochem. 73, 301–308.

    Article  PubMed  CAS  Google Scholar 

  21. Lew, R. A., Tetaz, T, and Smith, A. I. (1994) Characterization of a membrane-associated EC 3.4.24.15-like endopeptidase expressed by endothelial cells of the ovine median eminence. J. Neuroendocrinol. 6, 225–232.

    Article  PubMed  CAS  Google Scholar 

  22. Norman, M. U., Smith, A. I., and Lew, R. A. (1999) Role of calcium in the release of the bradykinin-degrading peptidase EC 3.4.24.16 from endothelial cells. Lett. Peptide Sci. 6, 349–352.

    CAS  Google Scholar 

  23. Lew, R. A., Boulos, E., Stewart, K. M., et al. (2000) Bradykinin analogs with β-amino acid substitutions reveal subtle differences in substrate specificity between the endopeptidases EC 3.4.24.15 and EC 3.4.24.16. J. Peptide Science 6, 440–445.

    Article  CAS  Google Scholar 

  24. Lew, R. A., Hey, N. J., Tetaz, T. J., Glucksman, M. J., Roberts, J. L., and Smith, A. I. (1995) Substrate specificity differences between recombinant rat testes endopeptidase EC 3.4.24.15 and the native brain enzyme. Biochem. Biophys. Res. Comm. 209, 788–795.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2004 Humana Press Inc.

About this protocol

Cite this protocol

Lew, R.A. (2004). HPLC in the Analysis of Peptide Metabolism. In: Aguilar, MI. (eds) HPLC of Peptides and Proteins. Methods in Molecular Biology™, vol 251. Springer, Totowa, NJ. https://doi.org/10.1385/1-59259-742-4:275

Download citation

  • DOI: https://doi.org/10.1385/1-59259-742-4:275

  • Publisher Name: Springer, Totowa, NJ

  • Print ISBN: 978-0-89603-977-3

  • Online ISBN: 978-1-59259-742-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics