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Fluorescent Substrates for Polyamine Catabolic Enzymes and Transport

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Polyamines

Part of the book series: Methods in Molecular Biology ((MIMB,volume 720))

Abstract

The most widely used methods for measuring polyamine enzyme activities are radioisotope methods that measure the radioactivity of compounds produced from radiolabeled substrate by the enzyme reaction. Several fluorescent polyamines have been developed for the measurement of the polyamine transport system (PTS) or transglutaminase. Although fluorophores in the fluorescent polyamines may affect the affinity of the polyamine moiety to the enzyme protein, the assays that use fluorescent substrate are sensitive and simple for common laboratory usage.

In this chapter, the uses of dansyl polyamines with a simple high-performance liquid chromatography system for the measurement of the PTS and polyamine catabolic enzymes including spermidine/spermine N1-acetyltransferase and N1-acetylpolyamine oxidase are described.

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References

  1. Morgan DML (1998) Polyamine Protocols. Methods in molecular biology, vol 79. Humana Press, Totowa

    Google Scholar 

  2. Halline AG, Brasitus TA (1990) Reversed phase high performance liquid chromatographic method for the measurement of polyamine oxidase activity. J Chromatogr 533:187–194

    Article  PubMed  CAS  Google Scholar 

  3. Baillon JG, Mamont PS, Wagner J, Gerhart R, Lux P (1988) Fluorinated analogues of spermidine as substrates of spermine synthase. Eur J Biochem 176:237–242

    Article  PubMed  CAS  Google Scholar 

  4. Vujcic S, Diegelman P, Bacchi C, Kramer D, Porter CW (2002) Identification and characterization of a novel flavin-containing spermine oxidase of mammalian cell origin. Biochem J 367:665–675

    Article  PubMed  CAS  Google Scholar 

  5. Xu YJ, Hara T, Samejima K, Sasaki H, Kobayashi M, Takahashi A, Niitsu M (2002) Simultaneous determination of endogenous and orally administered 15N-labeled polyamines in rat organs. Anal Biochem 301:255–260

    Article  PubMed  CAS  Google Scholar 

  6. Suzuki O, Matsumoto T, Katsumata Y (1984) Determination of polyamine oxidase activities in human tissues. Experimentia 40:838–839

    Article  CAS  Google Scholar 

  7. Enomoto K, Nagasaki T, Yamauchi A, Onoda J, Sakai K, Yoshida T, Maekawa K, Kinoshita Y, Nishino I, Kikuoka S, Fukunaga T, Kawamoto K, Numata Y, Takemoto H, Nagata K (2006) Development of high-throughput spermidine synthase activity assay using homogeneous time-resolved fluorescence. Anal Biochem 351:229–240

    Article  PubMed  CAS  Google Scholar 

  8. Aziz SM, Yatin M, Worthen DR, Lipke DW, Crooks PA (1998) A novel technique for visualizing the intracellular localization and distribution of transported polyamines in cultured pulmonary artery smooth muscle cells. J Pharm Biomed Anal 17:307–320

    Article  PubMed  CAS  Google Scholar 

  9. Cullis PM, Green RE, Merson-Davies L, Travis N (1999) Probing the mechanism of transport and compartmentalization of polyamines in mammalian cells. Chem Biol 6:717–729

    Article  PubMed  CAS  Google Scholar 

  10. Soulet D, Covassin L, Kaouass M, Charest-Gaudreault R, Audette M, Poulin R (2002) Role of endocytosis in the internalization of spermidine-C2-BODIPY, a highly fluorescent probe of polyamine transport. Biochem J 367:347–357

    Article  PubMed  CAS  Google Scholar 

  11. Phanstiel O IV, Kaur N, Delcros J-D (2007) Structure-activity investigations of polyamine-anthracene conjugates and their uptake via the polyamine transporter. Amino Acids 33:305–313

    Article  PubMed  CAS  Google Scholar 

  12. Guminski Y, Grousseaud M, Cugnasse S, Brel V, Annereau J-P, Vispe S, Guilbaud N, Barret J-M, Bailly C, Imbert T (2009) Synthesis of conjugated spermine derivatives with 7-nitrobenzoxazole (NBD), rhodamine and bodipy as new fluorescent probes for the polyamine transport system. Bioorg Med Chem Lett 19:2474–2477

    Article  PubMed  CAS  Google Scholar 

  13. Wu Y-W, Tsai Y-H (2006) A rapid transglutaminase assay for high throughput screening applications. J Biomol Screen 11:836–843

    Article  PubMed  CAS  Google Scholar 

  14. Takao K, Miyatake S, Fukazawa K, Wada M, Sugita Y, Shirahata A (2008) Measurement of spermidine/spermine-N1-acetyltransferase activity by high-performance liquid chromatography with N1-dansylnorspermine as the substrate. Anal Biochem 376:277–279

    Article  PubMed  CAS  Google Scholar 

  15. Takao K, Sugita Y, Shirahata A (2010) Assay of N1-acetylpolyamine oxidase with N1, N11-didansylnorspermine as the substrate by ion-pair reversed phase high performance liquid chromatography. Biol Pharm Bull 33:1089–1094

    Article  PubMed  CAS  Google Scholar 

  16. Takao K, Sugita Y, Shirahata A (2010) Evaluation method for polyamine uptake by N1-dansylspermine. Amino Acids 38:533–539

    Article  PubMed  CAS  Google Scholar 

  17. Holst CM, Nevsten P, Johansson F, Carlemalm E, Oredsson SM (2008) Subcellular distribution of spermidine/spermine N1-acetyltransferase. Cell Biol Int 32:39–47

    Article  PubMed  CAS  Google Scholar 

  18. Pegg AE (2008) Spermidine/spermine N1-acetyltransferase: a key metabolic regulator. Am J Physiol Endocrinol Metab 294:E995–E1010

    Article  PubMed  CAS  Google Scholar 

  19. Seiler N (1995) Polyamine oxidase, properties and functions. Prog Brain Res 106:333–344

    Article  PubMed  CAS  Google Scholar 

  20. Takao K, Shibata S, Ozawa T, Wada M, Sugita Y, Samejima K, Shirahata S (2009) A conceptual model of the polyamine binding site of N1-acetylpolyamine oxidase developed from a study of polyamine derivatives. Amino Acids 37:401–405

    Article  PubMed  CAS  Google Scholar 

  21. Seiler N, Dezeure F (1990) Polyamine transport in mammalian cells. Int J Biochem 22:211–218

    Article  PubMed  CAS  Google Scholar 

  22. Seiler N, Delcros JD, Molinoux JP (1996) Polyamine transport in mammalian cells. An update. Int J Biochem 28:843–861

    Article  CAS  Google Scholar 

  23. Bey P, Bolkenius FN, Seiler N, Casara P (1985) N-2, 3-butadienyl-1, 4-butanediamine derivatives: potent irreversible inactivators of mammalian polyamine oxidase. J Med Chem 28:1–2

    Article  PubMed  CAS  Google Scholar 

  24. Brunton VG, Grant HM, Wallace HM (1994) Spermine toxicity in BHL-21/C13 cells in the presence of bovine serum: the effect of aminoguanidine. Toxicol In Vitro 8:337–341

    Article  PubMed  CAS  Google Scholar 

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Correspondence to Akira Shirahata .

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Takao, K., Shirahata, A. (2011). Fluorescent Substrates for Polyamine Catabolic Enzymes and Transport. In: Pegg, A., Casero, Jr., R. (eds) Polyamines. Methods in Molecular Biology, vol 720. Humana Press. https://doi.org/10.1007/978-1-61779-034-8_30

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  • DOI: https://doi.org/10.1007/978-1-61779-034-8_30

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  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-033-1

  • Online ISBN: 978-1-61779-034-8

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