Advertisement

Polyamines pp 219-235 | Cite as

Identification and Assay of Allosteric Regulators of S-Adenosylmethionine Decarboxylase

  • Erin K. Willert
  • Lisa N. Kinch
  • Margaret A. Phillips
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 720)

Abstract

Polyamine biosynthesis is extensively regulated in cells by multiple mechanisms, including regulation of enzyme activity posttranslationally. The identified regulatory factors include both small molecules and regulatory proteins, and the mechanisms vary in different species across the evolutionary tree. Based on this diversity of mechanism, it is likely that regulatory factors of the pathway remain unidentified in many species. This article focuses on methods for identifying novel regulatory factors of polyamine biosynthesis as illustrated by the discovery of a novel protein activator of the key biosynthetic enzyme S-adenosylmethionine decarboxylase in the protozoan trypanosomatid parasites.

Key words

Trypanosoma Prozyme Polyamines Spermidine Putrescine Ornithine decarboxylase S-adenosylmethionine decarboxylase 

References

  1. 1.
    Pegg AE (2009) Mammalian polyamine metabolism and function. IUBMB Life 61:880–894PubMedCrossRefGoogle Scholar
  2. 2.
    Casero RA Jr, Marton LJ (2007) Targeting polyamine metabolism and function in cancer and other hyperproliferative diseases. Nat Rev Drug Discov 6:373–390PubMedCrossRefGoogle Scholar
  3. 3.
    Heby O, Persson L, Rentala M (2007) Targeting the polyamine biosynthetic enzymes: a promising approach to therapy of African sleeping sickness Chagas’ disease, and leishmaniasis. Amino Acids 33:359–366PubMedCrossRefGoogle Scholar
  4. 4.
    Priotto G, Kasparian S, Mutombo W, Ngouama D, Ghorashian S, Arnold U, Ghabri S, Baudin E, Buard V, Kazadi-Kyanza S, Ilunga M, Mutangala W, Pohlig G, Schmid C, Karunakara U, Torreele E, Kande V (2009) Nifurtimox-eflornithine combination therapy for second-stage African Trypanosoma brucei gambiense trypanosomiasis: a multicentre, randomised, phase III, non-inferiority trial. Lancet 374:56–64PubMedCrossRefGoogle Scholar
  5. 5.
    Casero RA, Pegg AE (2009) Polyamine catabolism and disease. Biochem J 421:323–338PubMedCrossRefGoogle Scholar
  6. 6.
    Bennett EM, Ekstrom JL, Pegg AE, Ealick SE (2002) Monomeric S-adenosylmethionine decarboxylase from plants provides an alternative to putrescine stimulation. Biochemistry 41:14509–14517PubMedCrossRefGoogle Scholar
  7. 7.
    Bale S, Lopez MM, Makhatadze GI, Fang Q, Pegg AE, Ealick SE (2008) Structural basis for putrescine activation of human S-adenosylmethionine decarboxylase. Bioche-mistry 47:13404–13417PubMedCrossRefGoogle Scholar
  8. 8.
    Ekstrom JL, Mathews II, Stanley BA, Pegg AE, Ealick SE (1999) The crystal structure of human S-adenosylmethionine decarboxylase at 2.25 A resolution reveals a novel fold. Structure 7:583–595PubMedCrossRefGoogle Scholar
  9. 9.
    Ekstrom JL, Tolbert WD, Xiong H, Pegg AE, Ealick SE (2001) Structure of a human S-adenosylmethionine decarboxylase self-­processing ester intermediate and mechanism of putrescine stimulation of processing as revealed by the H243A mutant. Biochemistry 40:9495–9504PubMedCrossRefGoogle Scholar
  10. 10.
    Willert EK, Fitzpatrick R, Phillips MA (2007) Allosteric regulation of an essential trypanosome polyamine biosynthetic enzyme by a catalytically dead homolog. Proc Natl Acad Sci U S A 104:8275–8280PubMedCrossRefGoogle Scholar
  11. 11.
    Willert EK, Phillips MA (2009) Cross-species activation of trypanosome S-adenosyl-methionine decarboxylase by the regulatory subunit prozyme. Mol Biochem Parasitol 168:1–6PubMedCrossRefGoogle Scholar
  12. 12.
    Beswick TC, Willert EK, Phillips MA (2006) Mechanisms of allosteric regulation of Trypanosoma cruzi S-adenosylmethionine decarboxylase. Biochemistry 45:7797–7807PubMedCrossRefGoogle Scholar
  13. 13.
    Willert EK, Phillips MA (2008) Regulated expression of an essential allosteric activator of polyamine biosynthesis in African trypanosomes. PLoS Pathog 4:e1000183PubMedCrossRefGoogle Scholar
  14. 14.
    Wu H, Min J, Zeng H, McCloskey DE, Ikeguchi Y, Loppnau P, Michael AJ, Pegg AE, Plotnikov AN (2008) Crystal structure of human spermine synthase: implications of substrate binding and catalytic mechanism. J Biol Chem 283:16135–16146PubMedCrossRefGoogle Scholar
  15. 15.
    Lee J, Sperandio V, Frantz DE, Longgood J, Camilli A, Phillips MA, Michael AJ (2009) An alternative polyamine biosynthetic pathway is widespread in bacteria and essential for biofilm formation in Vibrio cholerae. J Biol Chem 284:9899–9907PubMedCrossRefGoogle Scholar
  16. 16.
    Kinch LN, Scott JR, Ullman B, Phillips MA (1999) Cloning and kinetic characterization of the Trypanosoma cruzi S-adenosylmethionine decarboxylase. Mol Biochem Parasitol 101:1–11PubMedCrossRefGoogle Scholar
  17. 17.
    Lebowitz J, Lewis MS, Schuck P (2002) Modern analytical ultracentrifugation in protein science: a tutorial review. Protein Sci 11:2067–2079PubMedCrossRefGoogle Scholar
  18. 18.
    Osterman AL, Brooks HB, Jackson L, Abbott JJ, Phillips MA (1999) Lysine-69 plays a key role in catalysis by ornithine decarboxylase through acceleration of the Schiff base formation, decarboxylation, and product release steps. Biochemistry 38:11814–11826PubMedCrossRefGoogle Scholar
  19. 19.
    Copeland R (1996) Enzymes: a practical introduction to structure, mechanism, and data analysis. Wiley, New YorkGoogle Scholar
  20. 20.
    Stanley BA, Pegg AE (1991) Amino acid residues necessary for putrescine stimulation of human S-adenosylmethionine decarboxylase proenzyme processing and catalytic activity. J Biol Chem 266:18502–18506PubMedGoogle Scholar
  21. 21.
    Persson K, Aslund L, Grahn B, Hanke J, Heby O (1998) Trypanosoma cruzi has not last its s-adenosylmethionine decarboxylase: characterization of the gene and the encoded enzyme. Biochem J 333:527–537PubMedGoogle Scholar
  22. 22.
    Roberts S, Scott J, Gasteier J, Jiang Y, Brooks B, Jardim A, Carter N, Heby O, Ullman B (2002) S-adenosylmethionine decarboxylase from Leishmania donovani: molecular, genetic and biochemical characterization of null mutants and overproducers. J Biol Chem 277:5902–5909PubMedCrossRefGoogle Scholar
  23. 23.
    Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402PubMedCrossRefGoogle Scholar
  24. 24.
    Pei J, Kim BH, Grishin NV (2008) PROMALS3D: a tool for multiple protein sequence and structure alignments. Nucleic Acids Res 36:2295–2300PubMedCrossRefGoogle Scholar
  25. 25.
    Adachi J, Hasegawa M (1992) Molphy: programs for molecular phylogenetics based on maximum likelihood. Computer Science Monographs, Tokyo, Institute of Statistical Mathematics.Google Scholar
  26. 26.
    Osterman A, Grishin NV, Kinch LN, Phillips MA (1994) Formation of functional cross-species heterodimers of ornithine decarboxylase. Biochemistry 33:13662–13667PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Erin K. Willert
    • 1
  • Lisa N. Kinch
    • 2
  • Margaret A. Phillips
    • 1
  1. 1.Department of PharmacologyUniversity of Texas Southwestern Medical Center at DallasDallasUSA
  2. 2.Department of BiochemistryUniversity of Texas Southwestern Medical Center at DallasDallasUSA

Personalised recommendations