Plant Molecular Biology

, Volume 38, Issue 5, pp 725–734 | Cite as

The presence of CYP79 homologues in glucosinolate-producing plants shows evolutionary conservation of the enzymes in the conversion of amino acid to aldoxime in the biosynthesis of cyanogenic glucosides and glucosinolates

  • Søren Bak
  • Hanne Linde Nielsen
  • Barbara Ann Halkier
Article

Abstract

A cDNA encoding CYP79B1 has been isolated from Sinapis alba. CYP79B1 from S. alba shows 54% sequence identity and 73% similarity to sorghum CYP79A1 and 95% sequence identity to the Arabidopsis T42902, assigned CYP79B2. The high identity and similarity to sorghum CYP79A1, which catalyses the conversion of tyrosine to p-hydroxyphenylacetaldoxime in the biosynthesis of the cyanogenic glucoside dhurrin, suggests that CYP79B1 similarly catalyses the conversion of amino acid(s) to aldoxime(s) in the biosynthesis of glucosinolates. Within the highly conserved ‘PERF’ and the heme-binding region of A-type cytochromes, the CYP79 family has unique substitutions that define the family-specific consensus sequences of FXP(E/D)RH and SFSTG(K/R)RGC(A/I)A, respectively. Sequence analysis of PCR products generated with CYP79B subfamily-specific primers identified CYP79B homologues in Tropaeolum majus, Carica papaya, Arabidopsis, Brassica napus and S. alba. The five glucosinolate-producing plants identified a CYP79B amino acid consensus sequence KPERHLNECSEVTLTENDLRFISFSTGKRGC. The unique substitutions in the ‘PERF’ and the heme-binding domain and the high sequence identity and similarity of CYP79B1, CYP79B2 and CYP79A1, together with the isolation of CYP79B homologues in the distantly related Tropaeolaceae, Caricaceae and Brassicaceae within the Capparales order, show that the initial part of the biosynthetic pathway of glucosinolates and cyanogenic glucosides is catalysed by evolutionarily conserved cytochromes P450. This confirms that the appearance of glucosinolates in Capparales is based on a cyanogen ‘predisposition’. Identification of CYP79 homologues in glucosinolate-producing plants provides an important tool for tissue-specific regulation of the level of glucosinolates to improve nutritional value and pest resistance.

glucosinolates cyanogenic glucosides biosynthesis cytochrome P450 evolution Capparales 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Bak S, Kahn RA, Nielsen HL, Møller BL, Halkier BA: Cloning of three A-type cytochromes P450, CYP71E1, CYP98 and CYP99 from Sorghum bicolor (L.) Moench by a PCR approach and identification by expression in Escherichia coli of CYP71E1 as the oxime-metabolizing cytochrome P450 in the biosynthesis of the cyanogenic glucoside dhurrin. Plant Mol Biol 36: 393–405 (1998).Google Scholar
  2. 2.
    Bak S, Kahn RA, Olsen CE, Halkier BA: Cloning and expression in Escherichia coli of the obtusifoiol 14α-demethylase of Sorghum bicolor (L.) Moench, a cytochrome P450 orthologous to the sterol 14α-demethylases (CYP51) from fungi and mammals. Plant J 11: 191–201 (1997).Google Scholar
  3. 3.
    Barnes HJ: Maximizing expression of eukaryotic cytochrome P450s in Escherichia coli. Meth Enzymol 272: 3–14 (1996).Google Scholar
  4. 4.
    Barnes HJ, Arlotto MP, Waterman MR: Expression and enzymatic activity of recombinant cytochrome P450 17áhydroxylase in Escherichia coli. Proc Natl Acad Sci USA 88: 5597–5601 (1991).Google Scholar
  5. 5.
    Bennett RN, Donald A, Dawson GW, Hick AJ, Wallsgrove RM: Aldoxime-forming microsomal enzyme systems involved in the biosynthesis of glucosinolates in oilseed rape (Brassica napus) leaves. Plant Physiol 102: 1307–1312 (1993).Google Scholar
  6. 6.
    Bennett RN, Hick AJ, Dawson GW, Wallsgrove RM: Glucosinolate biosynthesis: further characterization of the aldoximeforming microsomal monooxygenases in oilseed rape leaves. Plant Physiol 109: 299–305 (1995).Google Scholar
  7. 7.
    Bennett RN, Kiddle G, Hick AJ, Dawson GW, Wallsgrove RM: Distribution and activity of microsomal NADPHdependent monooxygenases and amino acid decarboxylases in cruciferous content. Plant Cell Environ 19: 801–812 (1996).Google Scholar
  8. 8.
    Bennett RN, Kiddle G, Wallsgrove RM: Involvement of cytochrome P450 in glucosinolate biosynthesis in white mustard. A biochemical anomaly. Plant Physiol 114: 1283–1291 (1997).Google Scholar
  9. 9.
    Bennett RN, Kiddle G, Wallsgrove RM: Biosynthesis of benzylglucosinolate, cyanogenic glucosides and phenylpropanoids in Carica papaya. Phytochemistry 45: 59–66 (1997).Google Scholar
  10. 10.
    Conn EE: Cyanogenic Glycosides. In: Bell EA, Charlwood BV (eds) Secondary Plant Products, pp. 461–492. Springer-Verlag, Berlin/Heidelberg/ New York (1980).Google Scholar
  11. 11.
    Du L, Halkier BA: Isolation of a microsomal enzyme system involved in glucosinolate biosynthesis from seedlings of Tropaeolum majus L. Plant Physiol 111: 831–837 (1996).Google Scholar
  12. 12.
    Du L, Lykkefeldt J, Olsen CE, Halkier BA: Involvement of cytochrome P450 in oxime production in glucosinolate biosynthesis as demonstrated by an in vitro microsomal enzyme system isolated from jasmonic acid-induced seedlings of Sinapis alba L. Proc Natl Acad Sci USA 92: 12505–12509 (1995).Google Scholar
  13. 13.
    Durst F, Nelson DR: Diversity and evolution of plant P450 and P450-reductases. Drug Metabol Drug Interact 12: 189–206 (1995).Google Scholar
  14. 14.
    Ettlinger MG, Kjær A: Sulfur compounds in plants. Rec Adv Phytochem 1: 49–144 (1968).Google Scholar
  15. 15.
    Halkier BA: Catalytic reactivities and structure/function relationship of cytochrome P450 enzymes. Phytochemistry 43: 1–21 (1996).Google Scholar
  16. 16.
    Halkier BA: Glucosinolates. In: Ikan R (ed) Naturally Occurring Glycosides. John Wiley, New York (1998).Google Scholar
  17. 17.
    Halkier BA, Du L: The biosynthesis of glucosinolates. Trends Plant Sci 2: 425–431 (1997).Google Scholar
  18. 18.
    Halkier BA, Nielsen HL, Koch BM, Møller BL: Purification and characterization of recombinant cytochrome P450TYR expressed at high levels in Escherichia coli. Arch Biochem Biophys 322: 369–377 (1995).Google Scholar
  19. 19.
    Halkier BA, Sibbesen O, Koch B, Møller BL: Characterization of cytochrome P450tyr, a multifunctional heme-thiolate N-hydroxylase involved in the biosynthesis of the cyanogenic glucoside dhurrin. Drug Metabol Drug Interact 12: 285–297 (1995).Google Scholar
  20. 20.
    Hasemann CA, Kurumbali RG, Boddupalli SS, Peterson JA, Deisenhofer J: Structure and function of cytochrome P450: a comparative analysis of three crystal structures. Structure 2: 41–62 (1995).Google Scholar
  21. 21.
    Hogge RL, Reed DW, Underhill EW: HPLC separation of glucosinolates from leaves and seeds of Arabidopsis thaliana and their identification using thermospray liquid chromatographymass spectrometry. Chromatog Sci 26: 551–560 (1988).Google Scholar
  22. 22.
    Kahn RA, Bak S, Svendsen I, Halkier BA, Møller BL: Isolation and reconstitution of cytochrome P450ox and in vitro reconstitution of the entire biosynthetic pathway of the cyanogenic glucoside dhurrin from sorghum. Plant Physiol 115: 1661–1670 (1997).Google Scholar
  23. 23.
    Koch B, Nielsen VS, Halkier BA, Olsen CE, Møller BL: The biosynthesis of cyanogenic glucosides in seedlings of cassava (Manihot esculenta Crantz). Arch Biochem Biophys 292: 141–150 (1992).Google Scholar
  24. 24.
    Koch BA, Sibbesen O, Halkier BA, Svendsen I, Møller BL: The primary sequence of cytochrome P450tyr, the multifunctional N-hydroxylase catalyzing the conversion of L-tyrosine to p-hydroxyphenylacetaldehyde oxime in the biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor (L.) Moench. Arch Biochem Biophys 323: 177–186 (1995).Google Scholar
  25. 25.
    Ludwig-Muller J, Hilgenberg W: A plasma membrane-bound enzyme oxidizes L-tryptophan to indole-3-acetaldoxime. Physiol Plant 74: 240–250 (1988).Google Scholar
  26. 26.
    Poulton JE, Møller BL: Glucosinolates. Meth Plant Biochem 9: 209–237 (1993).Google Scholar
  27. 27.
    Rodman JE, Karol KG, Price RA, Sytsma KJ: Molecules, morphology and Dahlgren's expanded order Capparales. System Bot 21: 289–307 (1996).Google Scholar
  28. 28.
    Saupe SG: Cyanogenic compounds and angiosperm phylogeny. In: Young DA, Seigler DS (eds) Angiosperm Phylogeny, pp. 80–116. Praeger Scientific, New York (1981).Google Scholar
  29. 29.
    Schuler MA: Plant cytochrome P450 monooxygenases. Crit Rev Plant Sci 15: 235–284 (1996).Google Scholar
  30. 30.
    Sibbesen O, Koch B, Halkier BA, Møller BL: Isolation of the heme-thiolate enzyme cytochrome P-450TYR, which catalyses the committed step in the biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor (L.) Moench. Proc Natl Acad Sci USA 91: 9740–9744 (1994).Google Scholar
  31. 31.
    Sibbesen O, Koch B, Halkier BA, Møller BL: Cytochrome P450tyr is a multifunctional heme-thiolate enzyme catalyzing the conversion of L-tyrosine to p-hydroxyphenylacetaldehyde oxime in the biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor (L.) Moench. J Biol Chem 270: 3506–3511 (1995).Google Scholar
  32. 32.
    Sørensen H: Glucosinolates: structure, properties, function. In: Shahidi F (ed) Canola and Rapeseed, pp. 149–172. Van Nostrand Reinhold, New York (1991).Google Scholar

Copyright information

© Kluwer Academic Publishers 1998

Authors and Affiliations

  • Søren Bak
    • 1
  • Hanne Linde Nielsen
    • 1
  • Barbara Ann Halkier
    • 1
  1. 1.Plant Biochemistry Laboratory, Department of Plant BiologyThe Royal Veterinary and Agricultural UniversityFrederiksberg C, CopenhagenDenmark

Personalised recommendations