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. 1990 Oct 15;271(2):559–562. doi: 10.1042/bj2710559

Slow tight-binding inhibition of prolyl endopeptidase by benzyloxycarbonyl-prolyl-prolinal.

A V Bakker 1, S Jung 1, R W Spencer 1, F J Vinick 1, W S Faraci 1
PMCID: PMC1149592  PMID: 2241932

Abstract

Prolyl endopeptidase is a serine proteinase that specifically cleaves peptides on the carboxy side of proline residues. Wilk & Orlowski [(1983) J. Neurochem. 41, 69-75] have shown that benzyloxycarbonyl-prolyl-prolinal (Z-prolyl-prolinal) is a potent inhibitor of prolyl endopeptidase. We show that Z-prolyl-prolinal is a slow-binding inhibitor of mouse brain prolyl endopeptidase with Ki 0.35 +/- 0.05 nM. Kinetic analysis indicates that the mechanism is a simple, but slow, reversible equilibrium between free and bound enzyme (E + I in equilibrium EI) with rate constants for association (kon) and dissociation (koff) of 1.6 X 10(5) M-1.s-1 and approx. 4 X 10(-5) s-1 respectively. Slow-binding inhibition is dependent on the presence of the aldehyde group since the alcohol (Z-prolyl-prolinol) is a rapid and 50,000-fold poorer inhibitor (Ki 19 microM). Prolyl endopeptidase from human brain is also inhibited by Z-prolyl-prolinal with kinetics similar to those of the mouse brain enzyme.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cleland W. W. Statistical analysis of enzyme kinetic data. Methods Enzymol. 1979;63:103–138. doi: 10.1016/0076-6879(79)63008-2. [DOI] [PubMed] [Google Scholar]
  2. Copp L. J., Krantz A., Spencer R. W. Kinetics and mechanism of human leukocyte elastase inactivation by ynenol lactones. Biochemistry. 1987 Jan 13;26(1):169–178. doi: 10.1021/bi00375a024. [DOI] [PubMed] [Google Scholar]
  3. Erion M. D., Walsh C. T. 1-Aminocyclopropanephosphonate: time-dependent inactivation of 1-aminocyclopropanecarboxylate deaminase and Bacillus stearothermophilus alanine racemase by slow dissociation behavior. Biochemistry. 1987 Jun 16;26(12):3417–3425. doi: 10.1021/bi00386a025. [DOI] [PubMed] [Google Scholar]
  4. Friedman T. C., Orlowski M., Wilk S. Prolyl endopeptidase: inhibition in vivo by N-benzyloxycarbonyl-prolyl-prolinal. J Neurochem. 1984 Jan;42(1):237–241. doi: 10.1111/j.1471-4159.1984.tb09723.x. [DOI] [PubMed] [Google Scholar]
  5. Kennedy W. P., Schultz R. M. Mechanism of association of a specific aldehyde "transition-state analogue" to the active site of alpha-chymotrypsin. Biochemistry. 1979 Jan 23;18(2):349–356. doi: 10.1021/bi00569a019. [DOI] [PubMed] [Google Scholar]
  6. Morrison J. F., Walsh C. T. The behavior and significance of slow-binding enzyme inhibitors. Adv Enzymol Relat Areas Mol Biol. 1988;61:201–301. doi: 10.1002/9780470123072.ch5. [DOI] [PubMed] [Google Scholar]
  7. Orlowski M., Wilk E., Pearce S., Wilk S. Purification and properties of a prolyl endopeptidase from rabbit brain. J Neurochem. 1979 Aug;33(2):461–469. doi: 10.1111/j.1471-4159.1979.tb05176.x. [DOI] [PubMed] [Google Scholar]
  8. Stein R. L., Strimpler A. M., Edwards P. D., Lewis J. J., Mauger R. C., Schwartz J. A., Stein M. M., Trainor D. A., Wildonger R. A., Zottola M. A. Mechanism of slow-binding inhibition of human leukocyte elastase by trifluoromethyl ketones. Biochemistry. 1987 May 19;26(10):2682–2689. doi: 10.1021/bi00384a005. [DOI] [PubMed] [Google Scholar]
  9. Tsuru D., Yoshimoto T., Koriyama N., Furukawa S. Thiazolidine derivatives as potent inhibitors specific for prolyl endopeptidase. J Biochem. 1988 Oct;104(4):580–586. doi: 10.1093/oxfordjournals.jbchem.a122514. [DOI] [PubMed] [Google Scholar]
  10. Wilk S., Orlowski M. Inhibition of rabbit brain prolyl endopeptidase by n-benzyloxycarbonyl-prolyl-prolinal, a transition state aldehyde inhibitor. J Neurochem. 1983 Jul;41(1):69–75. doi: 10.1111/j.1471-4159.1983.tb11815.x. [DOI] [PubMed] [Google Scholar]
  11. Wilk S. Prolyl endopeptidase. Life Sci. 1983 Nov 28;33(22):2149–2157. doi: 10.1016/0024-3205(83)90285-0. [DOI] [PubMed] [Google Scholar]
  12. Yoshimoto T., Ogita K., Walter R., Koida M., Tsuru D. Post-proline cleaving enzyme. Synthesis of a new fluorogenic substrate and distribution of the endopeptidase in rat tissues and body fluids of man. Biochim Biophys Acta. 1979 Aug 15;569(2):184–192. doi: 10.1016/0005-2744(79)90053-6. [DOI] [PubMed] [Google Scholar]

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