Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera
Anthocyanidin reductase from Vitis vinifera catalyzes an NADPH-dependent double reduction of anthocyanidins. At pH 7.5 and 30 °C, steady-state kinetics support a hyperbolic and rapid-equilibrium ordered mechanism, with NADPH binding first, K M(cyan) = 2.82 ± 0.66 μM and K i(NADPH) = 111 ± 23 μM. The...
Gespeichert in:
| Veröffentlicht in: | Archives of biochemistry and biophysics 2009-11, Vol.491 (1), p.61-68 |
|---|---|
| Hauptverfasser: | , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 68 |
|---|---|
| container_issue | 1 |
| container_start_page | 61 |
| container_title | Archives of biochemistry and biophysics |
| container_volume | 491 |
| creator | Gargouri, Mahmoud Gallois, Bernard Chaudière, Jean |
| description | Anthocyanidin reductase from
Vitis vinifera catalyzes an NADPH-dependent double reduction of anthocyanidins. At pH 7.5 and 30
°C, steady-state kinetics support a hyperbolic and rapid-equilibrium ordered mechanism, with NADPH binding first,
K
M(cyan)
=
2.82
±
0.66
μM and
K
i(NADPH)
=
111
±
23
μM. The chromatographic method of Hummel and Dreyer was used for binding-equilibrium studies of NADPH, NADP
+ and catechin, at pH 7. This confirmed hyperbolic binding of NADPH and NADP
+ to the free enzyme, with a single binding site each and with dissociation constants
K
NADPH
=
45.9
±
2
μM and
K
NADP+
=
83
±
5
μM. There was no significant binding of catechin. We conclude (i) that the most likely mechanism is sequential ordered Bi Uni Uni Bi, with NADPH binding first and NADP
+ released last, and (ii) that internal conversion of the first ternary complex, i.e. that associated with the first hydride transfer, is rate-limiting. |
| doi_str_mv | 10.1016/j.abb.2009.09.010 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_734117412</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><els_id>S0003986109003087</els_id><sourcerecordid>734117412</sourcerecordid><originalsourceid>FETCH-LOGICAL-c352t-6c8d6ad0b95261766c9578c0ee236a4da8404b1de0690d492f6137e0e2fcabdf3</originalsourceid><addsrcrecordid>eNp9kE1Lw0AQhhdRbK3-AC-Sm6fUmU26SfCkxS8oeNFel83uRKc2ie4mhf57U1rwJgwMDM_7wjxCXCJMEVDdrKamLKcSoJjuBuFIjBEKFUOSp8diDABJXOQKR-IshBUAYqrkqRhhkWUyn8mxWN5z47j5iOmn5zWXnvs6Mo2Lvrihjm0Uut4xhaithnP32dqtaXhIRJ5cbzsTKKp8W0dL7jhEG264Im_OxUll1oEuDnsi3h8f3ubP8eL16WV-t4htMpNdrGzulHFQFjOpMFPKFrMst0AkE2VSZ_IU0hIdgSrApYWsFCYZAcnKmtJVyURc73u_ffvTU-h0zcHSem0aavugsyRFzFKUA4l70vo2BE-V_vZcG7_VCHpnU6_0YFPvbOrdIAyZq0N7X9bk_hIHfQNwuwdo-HHD5HWwTI0lx55sp13L_9T_Ap0OhgE</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>734117412</pqid></control><display><type>article</type><title>Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera</title><source>MEDLINE</source><source>Access via ScienceDirect (Elsevier)</source><creator>Gargouri, Mahmoud ; Gallois, Bernard ; Chaudière, Jean</creator><creatorcontrib>Gargouri, Mahmoud ; Gallois, Bernard ; Chaudière, Jean</creatorcontrib><description>Anthocyanidin reductase from
Vitis vinifera catalyzes an NADPH-dependent double reduction of anthocyanidins. At pH 7.5 and 30
°C, steady-state kinetics support a hyperbolic and rapid-equilibrium ordered mechanism, with NADPH binding first,
K
M(cyan)
=
2.82
±
0.66
μM and
K
i(NADPH)
=
111
±
23
μM. The chromatographic method of Hummel and Dreyer was used for binding-equilibrium studies of NADPH, NADP
+ and catechin, at pH 7. This confirmed hyperbolic binding of NADPH and NADP
+ to the free enzyme, with a single binding site each and with dissociation constants
K
NADPH
=
45.9
±
2
μM and
K
NADP+
=
83
±
5
μM. There was no significant binding of catechin. We conclude (i) that the most likely mechanism is sequential ordered Bi Uni Uni Bi, with NADPH binding first and NADP
+ released last, and (ii) that internal conversion of the first ternary complex, i.e. that associated with the first hydride transfer, is rate-limiting.</description><identifier>ISSN: 0003-9861</identifier><identifier>EISSN: 1096-0384</identifier><identifier>DOI: 10.1016/j.abb.2009.09.010</identifier><identifier>PMID: 19772852</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Anthocyanidin reductase ; Anthocyanins - metabolism ; Binding-equilibrium ; Catechin - metabolism ; Cyanidin ; Double reduction ; Hummel and Dreyer ; Hydrogen-Ion Concentration ; Kinetics ; Ligands ; NADH, NADPH Oxidoreductases - metabolism ; NADP - metabolism ; NADPH ; Oxygen - metabolism ; Protein Binding ; Spectrophotometry ; Steady-state kinetics ; Vitis - enzymology</subject><ispartof>Archives of biochemistry and biophysics, 2009-11, Vol.491 (1), p.61-68</ispartof><rights>2009 Elsevier Inc.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c352t-6c8d6ad0b95261766c9578c0ee236a4da8404b1de0690d492f6137e0e2fcabdf3</citedby><cites>FETCH-LOGICAL-c352t-6c8d6ad0b95261766c9578c0ee236a4da8404b1de0690d492f6137e0e2fcabdf3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.abb.2009.09.010$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19772852$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Gargouri, Mahmoud</creatorcontrib><creatorcontrib>Gallois, Bernard</creatorcontrib><creatorcontrib>Chaudière, Jean</creatorcontrib><title>Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera</title><title>Archives of biochemistry and biophysics</title><addtitle>Arch Biochem Biophys</addtitle><description>Anthocyanidin reductase from
Vitis vinifera catalyzes an NADPH-dependent double reduction of anthocyanidins. At pH 7.5 and 30
°C, steady-state kinetics support a hyperbolic and rapid-equilibrium ordered mechanism, with NADPH binding first,
K
M(cyan)
=
2.82
±
0.66
μM and
K
i(NADPH)
=
111
±
23
μM. The chromatographic method of Hummel and Dreyer was used for binding-equilibrium studies of NADPH, NADP
+ and catechin, at pH 7. This confirmed hyperbolic binding of NADPH and NADP
+ to the free enzyme, with a single binding site each and with dissociation constants
K
NADPH
=
45.9
±
2
μM and
K
NADP+
=
83
±
5
μM. There was no significant binding of catechin. We conclude (i) that the most likely mechanism is sequential ordered Bi Uni Uni Bi, with NADPH binding first and NADP
+ released last, and (ii) that internal conversion of the first ternary complex, i.e. that associated with the first hydride transfer, is rate-limiting.</description><subject>Anthocyanidin reductase</subject><subject>Anthocyanins - metabolism</subject><subject>Binding-equilibrium</subject><subject>Catechin - metabolism</subject><subject>Cyanidin</subject><subject>Double reduction</subject><subject>Hummel and Dreyer</subject><subject>Hydrogen-Ion Concentration</subject><subject>Kinetics</subject><subject>Ligands</subject><subject>NADH, NADPH Oxidoreductases - metabolism</subject><subject>NADP - metabolism</subject><subject>NADPH</subject><subject>Oxygen - metabolism</subject><subject>Protein Binding</subject><subject>Spectrophotometry</subject><subject>Steady-state kinetics</subject><subject>Vitis - enzymology</subject><issn>0003-9861</issn><issn>1096-0384</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kE1Lw0AQhhdRbK3-AC-Sm6fUmU26SfCkxS8oeNFel83uRKc2ie4mhf57U1rwJgwMDM_7wjxCXCJMEVDdrKamLKcSoJjuBuFIjBEKFUOSp8diDABJXOQKR-IshBUAYqrkqRhhkWUyn8mxWN5z47j5iOmn5zWXnvs6Mo2Lvrihjm0Uut4xhaithnP32dqtaXhIRJ5cbzsTKKp8W0dL7jhEG264Im_OxUll1oEuDnsi3h8f3ubP8eL16WV-t4htMpNdrGzulHFQFjOpMFPKFrMst0AkE2VSZ_IU0hIdgSrApYWsFCYZAcnKmtJVyURc73u_ffvTU-h0zcHSem0aavugsyRFzFKUA4l70vo2BE-V_vZcG7_VCHpnU6_0YFPvbOrdIAyZq0N7X9bk_hIHfQNwuwdo-HHD5HWwTI0lx55sp13L_9T_Ap0OhgE</recordid><startdate>20091101</startdate><enddate>20091101</enddate><creator>Gargouri, Mahmoud</creator><creator>Gallois, Bernard</creator><creator>Chaudière, Jean</creator><general>Elsevier Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20091101</creationdate><title>Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera</title><author>Gargouri, Mahmoud ; Gallois, Bernard ; Chaudière, Jean</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c352t-6c8d6ad0b95261766c9578c0ee236a4da8404b1de0690d492f6137e0e2fcabdf3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Anthocyanidin reductase</topic><topic>Anthocyanins - metabolism</topic><topic>Binding-equilibrium</topic><topic>Catechin - metabolism</topic><topic>Cyanidin</topic><topic>Double reduction</topic><topic>Hummel and Dreyer</topic><topic>Hydrogen-Ion Concentration</topic><topic>Kinetics</topic><topic>Ligands</topic><topic>NADH, NADPH Oxidoreductases - metabolism</topic><topic>NADP - metabolism</topic><topic>NADPH</topic><topic>Oxygen - metabolism</topic><topic>Protein Binding</topic><topic>Spectrophotometry</topic><topic>Steady-state kinetics</topic><topic>Vitis - enzymology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Gargouri, Mahmoud</creatorcontrib><creatorcontrib>Gallois, Bernard</creatorcontrib><creatorcontrib>Chaudière, Jean</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Archives of biochemistry and biophysics</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Gargouri, Mahmoud</au><au>Gallois, Bernard</au><au>Chaudière, Jean</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera</atitle><jtitle>Archives of biochemistry and biophysics</jtitle><addtitle>Arch Biochem Biophys</addtitle><date>2009-11-01</date><risdate>2009</risdate><volume>491</volume><issue>1</issue><spage>61</spage><epage>68</epage><pages>61-68</pages><issn>0003-9861</issn><eissn>1096-0384</eissn><abstract>Anthocyanidin reductase from
Vitis vinifera catalyzes an NADPH-dependent double reduction of anthocyanidins. At pH 7.5 and 30
°C, steady-state kinetics support a hyperbolic and rapid-equilibrium ordered mechanism, with NADPH binding first,
K
M(cyan)
=
2.82
±
0.66
μM and
K
i(NADPH)
=
111
±
23
μM. The chromatographic method of Hummel and Dreyer was used for binding-equilibrium studies of NADPH, NADP
+ and catechin, at pH 7. This confirmed hyperbolic binding of NADPH and NADP
+ to the free enzyme, with a single binding site each and with dissociation constants
K
NADPH
=
45.9
±
2
μM and
K
NADP+
=
83
±
5
μM. There was no significant binding of catechin. We conclude (i) that the most likely mechanism is sequential ordered Bi Uni Uni Bi, with NADPH binding first and NADP
+ released last, and (ii) that internal conversion of the first ternary complex, i.e. that associated with the first hydride transfer, is rate-limiting.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>19772852</pmid><doi>10.1016/j.abb.2009.09.010</doi><tpages>8</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0003-9861 |
| ispartof | Archives of biochemistry and biophysics, 2009-11, Vol.491 (1), p.61-68 |
| issn | 0003-9861 1096-0384 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_734117412 |
| source | MEDLINE; Access via ScienceDirect (Elsevier) |
| subjects | Anthocyanidin reductase Anthocyanins - metabolism Binding-equilibrium Catechin - metabolism Cyanidin Double reduction Hummel and Dreyer Hydrogen-Ion Concentration Kinetics Ligands NADH, NADPH Oxidoreductases - metabolism NADP - metabolism NADPH Oxygen - metabolism Protein Binding Spectrophotometry Steady-state kinetics Vitis - enzymology |
| title | Binding-equilibrium and kinetic studies of anthocyanidin reductase from Vitis vinifera |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-29T05%3A08%3A01IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_cross&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Binding-equilibrium%20and%20kinetic%20studies%20of%20anthocyanidin%20reductase%20from%20Vitis%20vinifera&rft.jtitle=Archives%20of%20biochemistry%20and%20biophysics&rft.au=Gargouri,%20Mahmoud&rft.date=2009-11-01&rft.volume=491&rft.issue=1&rft.spage=61&rft.epage=68&rft.pages=61-68&rft.issn=0003-9861&rft.eissn=1096-0384&rft_id=info:doi/10.1016/j.abb.2009.09.010&rft_dat=%3Cproquest_cross%3E734117412%3C/proquest_cross%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=734117412&rft_id=info:pmid/19772852&rft_els_id=S0003986109003087&rfr_iscdi=true |