Advertisement

Benzoate-CoA ligase contributes to the biosynthesis of biphenyl phytoalexins in elicitor-treated pear cell cultures

  • Shashank Sagar Saini
  • Mariam Gaid
  • Debabrata SircarEmail author
Original Article

Abstract

Key message

Benzoate-Coenzyme A ligase enzyme activity catalyzing the conversion of free benzoic acid to benzoyl-CoA was detected and biochemically characterized in the elicitor-treated pear cell cultures.

Abstract

Asian pear (Pyrus pyrifolia) is an economically and nutritionally important fruit-bearing tree of the subtribe Malinae. Upon pathogen attack, pears produce unique benzoate-derived biphenyl phytoalexins. The upstream biosynthesis of the biphenyl in Malinae is still incomplete. Previously, protein preparations from yeast extract-treated pear cultures were able to convert l-phenylalanine to cinnamic acid catalyzed by the activity of the phenylalanine ammonia lyase. The same extract was able to perform a C2 side-chain cleavage of cinnamic acid to benzaldehyde followed by oxidation of the latter to benzoic acid owing to the molecularly-undefined benzaldehyde synthase and benzaldehyde dehydrogenase activities, respectively. The biosynthesis of biphenyls starts with benzoate-Coenzyme A ligase (BZL), which converts benzoic acid to benzoyl-CoA. Subsequently, the previously-defined biphenyl synthase uses benzoyl-CoA to form the biphenyls. The current study reports the first time detection and characterization of BZL activity in elicitor-treated pear cell cultures. The preferred substrate was benzoic acid (Km = 62 ± 4 µM). Magnesium or manganese was prerequisite for the activity, which was enhanced by ~ 70% in the presence of potassium. Maximum BZL activity was observed 18 h post elicitation, which is in agreement with the coordinate induction reported for the enzymes in the same pathway. The induced BZL activity preceded the accumulation of biphenyls supporting its involvement in their biosynthesis.

Keywords

Benzoate-coenzyme A ligase Benzoic acid Biphenyl Cinnamate-CoA ligase Pyrus pyrifolia 

Notes

Acknowledgements

This research work was supported by a start up research grant (FIG 100624 to D. Sircar) from the Indian Institute of Technology Roorkee. SSS is thankful to MHRD-research assistantship (MHRD02-23-200-429) from Indian Institute of Technology Roorkee for perusing his doctoral studies. We thank Dr. Till Beuerle (Technische Universität Braunschweig, Germany) for ESI–MS analyses.

Author contribution statement

DS conceived and designed the study. SSS performed experiments, data processing and analyses. MG and DS performed data analyses and wrote the manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

References

  1. Abd El-Mawla AM, Beerhues L (2002) Benzoic acid biosynthesis in cell cultures of Hypericum androsaemum. Planta 214:727–733.  https://doi.org/10.1007/s004250100657 CrossRefPubMedGoogle Scholar
  2. Barillas W, Beerhues L (1997) 3-Hydroxybenzoate:coenzyme A ligase and 4-coumarate: coenzyme A ligase from cultured cells of Centaurium erythraea. Planta 202:112–116CrossRefGoogle Scholar
  3. Barillas W, Beerhues L (2000) 3-Hydroxybenzoate:coenzyme A ligase from cell cultures of Centaurium erythraea: isolation and characterization. Biol Chem 381:155–160CrossRefGoogle Scholar
  4. Beuerle T, Pichersky E (2002a) Enzymatic synthesis and purification of aromatic coenzyme A esters. Anal Biochem 302:305–312CrossRefGoogle Scholar
  5. Beuerle T, Pichersky E (2002b) Purification and characterization of benzoate:coenzyme A ligase from Clarkia breweri. Arch Biochem Biophys 400:258–264CrossRefGoogle Scholar
  6. Bjorklund JA, Leete E (1992) Biosynthesis of the benzoyl moiety of cocaine from cinnamic acid via (R)-(+)-3-hydroxy-3-phenylpropanoic acid. Phytochemistry 31:3883–3887.  https://doi.org/10.1016/S0031-9422(00)97546-0 CrossRefGoogle Scholar
  7. Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  8. Bussell JD, Reichelt M, Wiszniewski AAG et al (2014) Peroxisomal ATP-binding cassette transporter comatose and the multifunctional protein abnormal inflorescence meristem are required for the production of benzoylated metabolites in Arabidopsis seeds. Plant Physiol 164:48–54.  https://doi.org/10.1104/pp.113.229807 CrossRefPubMedGoogle Scholar
  9. Chizzali C, Beerhues L (2012) Phytoalexins of the pyrinae: biphenyls and dibenzofurans. Beilstein J Org Chem 8:613–620.  https://doi.org/10.3762/bjoc.8.68 CrossRefPubMedPubMedCentralGoogle Scholar
  10. Chizzali C, Khalil MNA, Beuerle T et al (2012) Formation of biphenyl and dibenzofuran phytoalexins in the transition zones of fire blight-infected stems of Malus domestica cv. “Holsteiner Cox” and Pyrus communis cv. “Conference”. Phytochemistry 77:179–185.  https://doi.org/10.1016/j.phytochem.2012.01.023 CrossRefPubMedGoogle Scholar
  11. Chizzali C, Swiddan AK, Abdelaziz S et al (2016) Expression of biphenyl synthase genes and formation of phytoalexin compounds in three fire blight-infected Pyrus communis cultivars. PLoS One 11:e0158713.  https://doi.org/10.1371/journal.pone.0158713 CrossRefPubMedPubMedCentralGoogle Scholar
  12. Colquhoun TA, Marciniak DM, Wedde AE et al (2012) A peroxisomally localized acyl-activating enzyme is required for volatile benzenoid formation in a Petunia × hybrida cv. “Mitchell Diploid” flower. J Exp Bot 63:4821–4833.  https://doi.org/10.1093/jxb/ers153 CrossRefPubMedPubMedCentralGoogle Scholar
  13. Fotirić Akšić MM, Dabić DČ, Gašić UM et al (2015) Polyphenolic profile of pear leaves with different resistance to Pear Psylla (Cacopsylla pyri). J Agric Food Chem 63:7476–7486.  https://doi.org/10.1021/acs.jafc.5b03394 CrossRefPubMedGoogle Scholar
  14. Gaid MM, Scharnhop H, Ramadan H et al (2011) 4-Coumarate:coA ligase family members from elicitor-treated Sorbus aucuparia cell cultures. J Plant Physiol 168:944–951.  https://doi.org/10.1016/j.jplph.2010.11.021 CrossRefPubMedGoogle Scholar
  15. Gaid MM, Sircar D, Muller A et al (2012) Cinnamate:coA ligase initiates the biosynthesis of a benzoate-derived xanthone phytoalexin in Hypericum calycinum Cell Cultures. Plant Physiol 160:1267–1280.  https://doi.org/10.1104/pp.112.204180 CrossRefPubMedPubMedCentralGoogle Scholar
  16. Hüttner C, Beuerle T, Scharnhop H, Ernst L, Beerhues L (2010) Differential effect of elicitors on biphenyl and dibenzofuran formation in sorbus aucuparia cell cultures. J Agric Food Chem 58:11977-11984.CrossRefGoogle Scholar
  17. Jiang S, Zheng X, Yu P et al (2016) Primitive genepools of asian pears and their complex hybrid origins inferred from fluorescent sequence-specific amplification polymorphism (SSAP) markers based on LTR retrotransposons. PLoS One 11:e0149192.  https://doi.org/10.1371/journal.pone.0149192 CrossRefPubMedPubMedCentralGoogle Scholar
  18. Khalil MNA, Brandt W, Beuerle T et al (2015) O-Methyltransferases involved in biphenyl and dibenzofuran biosynthesis. Plant J 83:263–276.  https://doi.org/10.1111/tpj.12885 CrossRefPubMedGoogle Scholar
  19. Klempien A, Kaminaga Y, Qualley A et al (2012) Contribution of CoA ligases to benzenoid biosynthesis in Petunia flowers. Plant Cell 24:2015–2030.  https://doi.org/10.1105/tpc.112.097519 CrossRefPubMedPubMedCentralGoogle Scholar
  20. Liu B, Beuerle T, Klundt T, Beerhues L (2004) Biphenyl synthase from yeast-extract-treated cell cultures of Sorbus aucuparia. Planta 218:492–496.  https://doi.org/10.1007/s00425-003-1144-y CrossRefPubMedGoogle Scholar
  21. Qualley AV, Widhalm JR, Adebesin F et al (2012) Completion of the core -oxidative pathway of benzoic acid biosynthesis in plants. Proc Natl Acad Sci 109:16383–16388.  https://doi.org/10.1073/pnas.1211001109 CrossRefPubMedGoogle Scholar
  22. Saini SS, Teotia D, Gaid M et al (2017) Benzaldehyde dehydrogenase-driven phytoalexin biosynthesis in elicitor-treated Pyrus pyrifolia cell cultures. J Plant Physiol 215:154–162.  https://doi.org/10.1016/j.jplph.2017.06.004 CrossRefPubMedGoogle Scholar
  23. Saini SS, Teotia D, Gaid M, Sircar D (2019) A new enzymatic activity from elicitor-treated pear cell cultures converting trans-cinnamic acid to benzaldehyde. Physiol Plant 167:64–74.  https://doi.org/10.1111/ppl.12871 CrossRefPubMedGoogle Scholar
  24. Sarkate A, Banerjee S, Mir JI, Roy P, Sircar D (2017) Antioxidant and cytotoxic activity of bioactive phenolic metabolites isolated from the yeast-extract treated cell culture of apple. Plant Cell, Tissue Organ Cult 130:641–664CrossRefGoogle Scholar
  25. Sarkate A, Saini SS, Teotia D, Gaid M, Mir JI, Roy P, Agrawal PK, Sircar D (2018) Comparative metabolomics of scab-resistance and susceptible apple cell cultures in response to scab fungus elicitor treatment. Sci Reoprts 8:17844Google Scholar
  26. Sarkate A, Saini SS, Gaid M, Teotia D, Mir JI, Agrawal PK, Beerhues L, Sircar D (2019) Molecular cloning and functional analysis of a biphenyl phytoalexin-specific O-methyltransferase from apple cell suspension cultures. Planta 249:677.  https://doi.org/10.1007/s00425-018-3031-6 CrossRefPubMedGoogle Scholar
  27. Schmelz S, Naismith JH (2009) Adenylate-forming enzymes. Curr Opin Struct Biol 19:666–671CrossRefGoogle Scholar
  28. Sircar D, Gaid M, Chizzali C et al (2015) Biphenyl 4-hydroxylases involved in aucuparin biosynthesis in Rowan and Apple are CYP736A proteins. Plant Physiol 168:428–442.  https://doi.org/10.1104/pp.15.00074 CrossRefPubMedPubMedCentralGoogle Scholar
  29. Stewart CJ, Woods K, Macias G, Allan AC, Hellens RP, Noel JP (2017) Molecular architectures of benzoic acid-specific type III polyketide synthases. Acta Crystallogr D Struct Biol 73:1007–1019.  https://doi.org/10.1107/S2059798317016618 CrossRefPubMedPubMedCentralGoogle Scholar
  30. Stuible HP, Büttner D, Ehlting J et al (2000) Mutational analysis of 4-coumarate:coA ligase identifies functionally important amino acids and verifies its close relationship to other adenylate-forming enzymes. FEBS Lett 467:117–122.  https://doi.org/10.1016/S0014-5793(00)01133-9 CrossRefPubMedGoogle Scholar
  31. Teng Y, Tanabe K, Tamura F, Itai A (2002) Genetic relationships of Pyrus species and cultivars native to east asia revealed by randomly amplified polymorphic DNA markers. J Am Soc Hortic Sci 127:262–270.  https://doi.org/10.21273/jashs.127.2.262 CrossRefGoogle Scholar
  32. Teotia D, Gaid M, Saini SS et al (2019) Cinnamate-CoA ligase is involved in biosynthesis of benzoate-derived biphenyl phytoalexin in Malus domestica ‘Golden Delicious’ cell cultures. Plant J.  https://doi.org/10.1111/tpj.14506 CrossRefPubMedGoogle Scholar
  33. Van Moerkercke A, Schauvinhold I, Pichersky E et al (2009) A plant thiolase involved in benzoic acid biosynthesis and volatile benzenoid production. Plant J 60:292–302.  https://doi.org/10.1111/j.1365-313X.2009.03953.x CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Plant Molecular Biology Group, Biotechnology DepartmentIndian Institute of Technology RoorkeeRoorkeeIndia
  2. 2.Technische Universität Braunschweig, Institute of Pharmaceutical BiologyBraunschweigGermany
  3. 3.Department of Bioorganic ChemistryLeibniz Institute of Plant BiochemistryHalle (Saale)Germany

Personalised recommendations