Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity

Mechanical wounding and jasmonic acid (JA) treatment have been shown to be important factors in controlling laticifer differentiation in Hevea brasiliensis (rubber tree). With the long-term aim of potentially modifying the endogenous levels of JA in H. brasiliensis by gene transfer, we describe in t...

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Main Authors:	Norton, Gareth, Pappusamy, Arokiaraj, Yusof, Faridah, Pujade-Renaud, Valerie, Perkins, Mark, Griffiths, David, Jones, Heddwyn
Format:	Article
Language:	English
Published:	Elsevier 2007
Subjects:	Q Science (General)
Online Access:	http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/1/P_14.pdf

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spelling	iium-246592012-08-27T07:24:41Z http://irep.iium.edu.my/24659/ Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity Norton, Gareth Pappusamy, Arokiaraj Yusof, Faridah Pujade-Renaud, Valerie Perkins, Mark Griffiths, David Jones, Heddwyn Q Science (General) Mechanical wounding and jasmonic acid (JA) treatment have been shown to be important factors in controlling laticifer differentiation in Hevea brasiliensis (rubber tree). With the long-term aim of potentially modifying the endogenous levels of JA in H. brasiliensis by gene transfer, we describe in this paper the molecular cloning of a H. brasiliensis allene oxide synthase (AOS) cDNA and biochemical characterisation of the recombinant AOS (His6-HbAOS) enzyme. The AOS cDNA encodes a protein with the expected motifs present in CYP74A sub-group of the cytochrome P450 super-family of enzymes that metabolise 13-hydroperoxylinolenic acid (13-HPOT), the intermediate involved in JA synthesis. The recombinant H. brasiliensis AOS enzyme was estimated to have a high binding affinity for 13-HPOT with a Km value of 4.02 ± 0.64 μM. Consistent with previous studies, mammalian cycloxygenase (COX) and lipoxygenase (LOX) inhibitors were shown to significantly reduce His6-HbAOS enzyme activity. Although JA had no effect on His6-HbAOS, salicylic acid (SA) was shown to significantly inhibit the recombinant AOS enzyme activity in a dose dependent manner. Moreover, it was demonstrated that SA, and various analogues of SA, acted as competitive inhibitors of His6-HbAOS when 13-HPOT was used as substrate. We speculate that this effect of salicylates on AOS activity may be important in cross-talking between the SA and JA signalling pathways in plants during biotic/abiotic stress. Elsevier 2007-02 Article PeerReviewed application/pdf en http://irep.iium.edu.my/24659/1/P_14.pdf Norton, Gareth and Pappusamy, Arokiaraj and Yusof, Faridah and Pujade-Renaud, Valerie and Perkins, Mark and Griffiths, David and Jones, Heddwyn (2007) Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity. Plant Physiology and Biochemistry, 45 (2). pp. 129-138. ISSN 0981-942 http://www.sciencedirect.com/science/article/pii/S098194280700006X 10.1016/j.plaphy.2007.01.003
repository_type	Digital Repository
institution_category	Local University
institution	International Islamic University Malaysia
building	IIUM Repository
collection	Online Access
language	English
topic	Q Science (General)
spellingShingle	Q Science (General) Norton, Gareth Pappusamy, Arokiaraj Yusof, Faridah Pujade-Renaud, Valerie Perkins, Mark Griffiths, David Jones, Heddwyn Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
description	Mechanical wounding and jasmonic acid (JA) treatment have been shown to be important factors in controlling laticifer differentiation in Hevea brasiliensis (rubber tree). With the long-term aim of potentially modifying the endogenous levels of JA in H. brasiliensis by gene transfer, we describe in this paper the molecular cloning of a H. brasiliensis allene oxide synthase (AOS) cDNA and biochemical characterisation of the recombinant AOS (His6-HbAOS) enzyme. The AOS cDNA encodes a protein with the expected motifs present in CYP74A sub-group of the cytochrome P450 super-family of enzymes that metabolise 13-hydroperoxylinolenic acid (13-HPOT), the intermediate involved in JA synthesis. The recombinant H. brasiliensis AOS enzyme was estimated to have a high binding affinity for 13-HPOT with a Km value of 4.02 ± 0.64 μM. Consistent with previous studies, mammalian cycloxygenase (COX) and lipoxygenase (LOX) inhibitors were shown to significantly reduce His6-HbAOS enzyme activity. Although JA had no effect on His6-HbAOS, salicylic acid (SA) was shown to significantly inhibit the recombinant AOS enzyme activity in a dose dependent manner. Moreover, it was demonstrated that SA, and various analogues of SA, acted as competitive inhibitors of His6-HbAOS when 13-HPOT was used as substrate. We speculate that this effect of salicylates on AOS activity may be important in cross-talking between the SA and JA signalling pathways in plants during biotic/abiotic stress.
format	Article
author	Norton, Gareth Pappusamy, Arokiaraj Yusof, Faridah Pujade-Renaud, Valerie Perkins, Mark Griffiths, David Jones, Heddwyn
author_facet	Norton, Gareth Pappusamy, Arokiaraj Yusof, Faridah Pujade-Renaud, Valerie Perkins, Mark Griffiths, David Jones, Heddwyn
author_sort	Norton, Gareth
title	Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
title_short	Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
title_full	Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
title_fullStr	Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
title_full_unstemmed	Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
title_sort	characterisation of recombinant hevea brasiliensis allene oxide synthase: effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity
publisher	Elsevier
publishDate	2007
url	http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/ http://irep.iium.edu.my/24659/1/P_14.pdf
first_indexed	2023-09-18T20:36:58Z
last_indexed	2023-09-18T20:36:58Z
_version_	1777409110303571968

Characterisation of recombinant Hevea brasiliensis allene oxide synthase: Effects of cycloxygenase inhibitors, lipoxygenase inhibitors and salicyclate on enzyme activity

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