Cysteine is the presumed catalytic residue of Citrus sinensis phospholipid hydroperoxide glutathione peroxidase over‐expressed under salt stress
Identifieur interne : 000218 ( France/Analysis ); précédent : 000217; suivant : 000219Cysteine is the presumed catalytic residue of Citrus sinensis phospholipid hydroperoxide glutathione peroxidase over‐expressed under salt stress
Auteurs : Zehava Faltin [France] ; Luc Camoin [France] ; Gozal Ben-Hayyim [France] ; Avi Perl [France] ; Tal Beeor-Tzahar [France] ; Arthur Donny Strosberg [France] ; Doron Holland [France] ; Yuval Eshdat [France]Source :
- Physiologia Plantarum [ 0031-9317 ] ; 1998-12.
Abstract
A citrus salt‐stress associated protein (Cit‐SAP), partially purified from citrus cultured cells, was previously identified as the first plant phospholipid hydroperoxide glutathione peroxidase (PHGPx). The nucleotide sequence of its isolated gene (csa) revealed that a TGT, known as codon for Cys, encodes the presumed catalytic residue 41 in the polypeptide chain of Cit‐SAP. In animals, a TGA encodes the rare amino acid selenocysteine (Sec) as the catalytic residue of the analogous enzyme. It is of interest to establish whether the TGT codon for this catalytic residue in the plant enzyme is indeed translated to Cys and not to Sec, and to demonstrate the effect of such a change, if it exists, on the nature of the enzymatic activity of the plant enzyme as compared to that of the animal. In the present study, we have purified for the first time, by affinity chromatography, enzymatically active citrus PHGPx from recombinant Escherichia coli bearing the csa gene. Tryptic digestion of the purified enzyme followed by HPLC afforded the isolation of a peptide which contains residue 41, and its sequence analysis revealed that this residue is indeed a Cys, and not Sec. The enzymatic activity and specificity of the recombinant Cit‐SAP was found to be similar to that observed before for the partially purified plant enzyme. However, the rate of this activity was much lower towards phospholipid hydroperoxides, and none towards hydrogen peroxide, as compared to that of the animal analogue. It is therefore suggested that the presence of a Cys, and not Sec, as the catalytic residue in the plant enzyme, affects its enzymatic activity and may determine a different biological role for the plant PHGPx from that of the animal.
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DOI: 10.1034/j.1399-3054.1998.1040432.x
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Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Camoin, Luc" sort="Camoin, Luc" uniqKey="Camoin L" first="Luc" last="Camoin">Luc Camoin</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Ben Ayyim, Gozal" sort="Ben Ayyim, Gozal" uniqKey="Ben Ayyim G" first="Gozal" last="Ben-Hayyim">Gozal Ben-Hayyim</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Perl, Avi" sort="Perl, Avi" uniqKey="Perl A" first="Avi" last="Perl">Avi Perl</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Beeor Zahar, Tal" sort="Beeor Zahar, Tal" uniqKey="Beeor Zahar T" first="Tal" last="Beeor-Tzahar">Tal Beeor-Tzahar</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Strosberg, Arthur Donny" sort="Strosberg, Arthur Donny" uniqKey="Strosberg A" first="Arthur Donny" last="Strosberg">Arthur Donny Strosberg</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Holland, Doron" sort="Holland, Doron" uniqKey="Holland D" first="Doron" last="Holland">Doron Holland</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author><author><name sortKey="Eshdat, Yuval" sort="Eshdat, Yuval" uniqKey="Eshdat Y" first="Yuval" last="Eshdat">Yuval Eshdat</name><affiliation wicri:level="3"><country xml:lang="fr">France</country><wicri:regionArea>a Z. Faltin, G. Ben‐Hayyim, A. Perl, T. Beeor‐Tzahar, D. Holland and Y. Eshdat (corresponding author, email vayuval@volcani.agri.gov.il), Dept of Fruit Tree Breeding and Molecular Genetics, Agricultural Research Organization, P.O. Box 6, The Volcani Center, 50250 Bet‐Dagan, Israel; bL. Camoin and A. D. Strosberg, Inst Conchin de Génétique Moléculaire, CNRS UPR 415, 22 Rue Méchain, F‐75014 Paris</wicri:regionArea><placeName><region type="region" nuts="2">Île-de-France</region><settlement type="city">Paris</settlement></placeName></affiliation></author></analytic><monogr></monogr><series><title level="j">Physiologia Plantarum</title><idno type="ISSN">0031-9317</idno><idno type="eISSN">1399-3054</idno><imprint><publisher>Munksgaard International Publishers</publisher><pubPlace>Copenhagen</pubPlace><date type="published" when="1998-12">1998-12</date><biblScope unit="volume">104</biblScope><biblScope unit="issue">4</biblScope><biblScope unit="page" from="741">741</biblScope><biblScope unit="page" to="746">746</biblScope></imprint><idno type="ISSN">0031-9317</idno></series><idno type="istex">43E71C4413ED51A3DAA64020202D7C4B92E76666</idno><idno type="DOI">10.1034/j.1399-3054.1998.1040432.x</idno><idno type="ArticleID">PPL1040432</idno></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0031-9317</idno></seriesStmt></fileDesc><profileDesc><textClass></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">A citrus salt‐stress associated protein (Cit‐SAP), partially purified from citrus cultured cells, was previously identified as the first plant phospholipid hydroperoxide glutathione peroxidase (PHGPx). The nucleotide sequence of its isolated gene (csa) revealed that a TGT, known as codon for Cys, encodes the presumed catalytic residue 41 in the polypeptide chain of Cit‐SAP. In animals, a TGA encodes the rare amino acid selenocysteine (Sec) as the catalytic residue of the analogous enzyme. It is of interest to establish whether the TGT codon for this catalytic residue in the plant enzyme is indeed translated to Cys and not to Sec, and to demonstrate the effect of such a change, if it exists, on the nature of the enzymatic activity of the plant enzyme as compared to that of the animal. In the present study, we have purified for the first time, by affinity chromatography, enzymatically active citrus PHGPx from recombinant Escherichia coli bearing the csa gene. Tryptic digestion of the purified enzyme followed by HPLC afforded the isolation of a peptide which contains residue 41, and its sequence analysis revealed that this residue is indeed a Cys, and not Sec. The enzymatic activity and specificity of the recombinant Cit‐SAP was found to be similar to that observed before for the partially purified plant enzyme. However, the rate of this activity was much lower towards phospholipid hydroperoxides, and none towards hydrogen peroxide, as compared to that of the animal analogue. It is therefore suggested that the presence of a Cys, and not Sec, as the catalytic residue in the plant enzyme, affects its enzymatic activity and may determine a different biological role for the plant PHGPx from that of the animal.</div></front></TEI><affiliations><list><country><li>France</li></country><region><li>Île-de-France</li></region><settlement><li>Paris</li></settlement></list><tree><country name="France"><region name="Île-de-France"><name sortKey="Faltin, Zehava" sort="Faltin, Zehava" uniqKey="Faltin Z" first="Zehava" last="Faltin">Zehava Faltin</name></region><name sortKey="Beeor Zahar, Tal" sort="Beeor Zahar, Tal" uniqKey="Beeor Zahar T" first="Tal" last="Beeor-Tzahar">Tal Beeor-Tzahar</name><name sortKey="Ben Ayyim, Gozal" sort="Ben Ayyim, Gozal" uniqKey="Ben Ayyim G" first="Gozal" last="Ben-Hayyim">Gozal Ben-Hayyim</name><name sortKey="Camoin, Luc" sort="Camoin, Luc" uniqKey="Camoin L" first="Luc" last="Camoin">Luc Camoin</name><name sortKey="Eshdat, Yuval" sort="Eshdat, Yuval" uniqKey="Eshdat Y" first="Yuval" last="Eshdat">Yuval Eshdat</name><name sortKey="Holland, Doron" sort="Holland, Doron" uniqKey="Holland D" first="Doron" last="Holland">Doron Holland</name><name sortKey="Perl, Avi" sort="Perl, Avi" uniqKey="Perl A" first="Avi" last="Perl">Avi Perl</name><name sortKey="Strosberg, Arthur Donny" sort="Strosberg, Arthur Donny" uniqKey="Strosberg A" first="Arthur Donny" last="Strosberg">Arthur Donny Strosberg</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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